
LIBRARY OF CONGRESS, 

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('imp. _ Sh ftlf K2_ 




PRESENTED BY 



























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publications 

OF THE 

^University of Pennsylvania 


Contributions 

(W iJLrthu FR0M THE 

ZOOLOGICAL LABORATORY 

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FOR THE YEAR 

I899 


Comparative Cytological Studies with Especial 
Regard to the Morphology of 
the Nucleolus 


BY 

Thomas H. Montgomery, Jr., Ph. D. 

Assistant Professor of Zodlogy in the University of Pennsylvania 


Reprinted from Journal of Morphology, Vol. XV, No. 2 


Ginn & Co., Selling Agents, Boston, Mass. 
1900 

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COMPARATIVE CYTOLOGICAL STUDIES 
WITH ESPECIAL REGARD TO THE 
MORPHOLOGY OP THE 
NUCLEOLUS 


THOS. H. MONTGOMERY, JR., Ph.D. 

(lecturer in zoology, university of Pennsylvania) 


From the Laboratory of the 
Wistar Institute of Anatomy and Biology, Philadelphia 


r 


Reprinted from Journal of Morphology, Vol. XV, No. 2, 1898 








| v \ J 




BOSTON, U.S.A. 

GINN & COMPANY, PUBLISHERS 

Cbe gtljenarttm press 

1898 








COMPARATIVE CYTOLOGICAL STUDIES, WITH 
ESPECIAL REGARD TO THE MORPHOLOGY 
OF THE NUCLEOLUS. 

THOS. H. MONTGOMERY, Jr., Ph.D. 

(Lecturer in Zoology, University of Pennsylvania.) 

From the Laboratory of the Wistar Institute of Anatomy and Biology, 

Philadelphia. 


CONTENTS. 

PAGE 

I. Introduction. 266 

II. Review of the Literature upon Nucleoli. 267 

A. Zoological Literature. 268 

B. Botanical Literature. 375 

C. Synonyms of the Term Nucleolus. 399 

III. Observations. 400 

A . Methods of Study. 400 

B. Protozoa. 402 

1. Gregarine from Lineus gesserensis. 402 

2. Gregarine from Carinella annulata. 406 

C. Metazoa. 410 

a. Egg Cells. 410 

1. Montagua pilata (Verr.). 410 

2. Doto.;. 418 

3. Amphiporus glutinosus (Verr.). 418 

4. Tetrastemma catenulatum (Verr.) Mont. 423 

5. Tetrastemma elegans (Verr.). 431 

6. Zygonemertes virescens (Verr.) Montg. 433 

7. Stichostemma eilhardi (Montg.). 437 

8. Lineus gesserensis (O. F. M.). 446 

9. Siphonophore (Rodalia ?) . 451 

10. Polydora. 455 

11. Piscicola rapax (Verr.). 464 

b. Somatic Cells. 472 

12. Ganglion Cells of Doto. 472 

13. Ganglion Cells of Montagua pilata (Verr.). 473 

14. Ganglion Cells of Piscicola rapax (Verr.). 475 

15. Muscle Cells of Lineus gesserensis (O. F. M.). 475 

16. Muscle Cells of Piscicola rapax (Verr.). 477 

17. Blood Corpuscles of Doto. 478 

18. Giant Cells of Doto. 480 


































MONTGOMERY. 


266 


[VOL. XV. 


PAGE 


19. Gland cells of Piscicola rapax (Verr.). 483 

20. Mesenchym Cells of Cerebratulus lacteus (Verr.). 494 

21. Ganglion Cells of Nemerteans. 49^ 

IV. General Comparisons and Conclusions . 497 

Appendix to the Literature Reviews . 539 

Literature List. 542 

Explanation of Plates . 561 


I. INTRODUCTION. 

The following studies are based upon animal cells, both egg 
cells and somatic cells having been investigated. They were 
made, primarily, with a regard to the morphology of the true 
nucleoli (plasmosomes), though numerous other points in onto¬ 
genetic cellular development have been considered. In con¬ 
nection with these observations the zoological literature upon 
the subject of nucleoli has been reviewed as thoroughly as 
possible, and, less completely, the literature from the botan¬ 
ical standpoint as well; reviews are given of these observa¬ 
tions of previous writers. No attempt has been made to 
review the literature from the pathological standpoint. Under 
the caption “ General Comparisons and Conclusions ” are com¬ 
pared together the more important deductions from my own 
observations, and these are compared with those of previous 
investigators. 

The nucleoli are cellular structures which have been studied 
to much less extent than other constituents of the cell, and 
though there are numerous observations upon them, these are 
so scattered through works of more general import that it is 
well-nigh impossible to collect together all the previous inves¬ 
tigations upon the subject. I hope that this explanation may 
be taken as an apology by any authors whose papers I have 
chanced to overlook. 

At the laboratory of the Fish Commission at Woods Holl, 
the following species were collected by me : Montagna , Amphi- 
porus glntinosus, Tetrastemma catennlatum y Zygonemertes , 
Linens , Polydora , and Piscicola. At Sea Isle City, at the labo¬ 
ratory of the University of Pennsylvania : Tetrastemma elegans y 
DotOy and certain of the species found at the former locality. 
Stic hostemma was collected in the aquaria of the University 









No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 267 


of Berlin; and the preparations of the siphonophore Rodalia 
were kindly placed at my disposal by Dr. E. G. Conklin. 

Doto and Montagna belong to the family of the Aeolidiidae; 
AmphiporuSy Tetrastemma , Zygonemertes , and Stichostemma are 
Metanemertini; and Linens and Cerebratulus are Heteronemer- 
tini; Polydora is a Polychaete; and Piscicola a rhynchobdellid 
leech. 

The present paper was sent to Dr. Whitman, editor of 
the Journal of Morphology , on Feb. 3, 1897; on receiving the 
MSS. again in March, 1898, I was able to incorporate in the 
text reviews of the literature of the whole year 1897. No 
other changes of importance, however, were then made in the 
original text, except brief mention of observations which I had 
made in the past year. It is my intention to follow this paper 
by others on nucleolar structures, particularly on structures 
which have received but little consideration in the present 
paper, namely, “ double ” nucleoli and chromatin nucleoli. 

II. REVIEW OF THE LITERATURE UPON NUCLEOLI. 

In this review shall be considered separately, first, those 
papers from the zoological, and, second, those from the 
botanical standpoint. The references from zoological papers 
I have endeavored to make as complete as possible, while my 
citations from the observations of botanical observers are 
much less numerous, though even in this I have consulted the 
more important papers from 1880 to the present time. In 
referring to the zoological papers, I have taken them up in 
chronological order; and in doing so, shall treat separately 
the periods 1781-1860, 1861-69, 1870-79, and from the year 
1880 to the present time I shall treat the literature for each 
year separately, in order that the reader may more conven¬ 
iently be able to turn to the citations from a given paper. 
Under each year papers are reviewed according to the alpha¬ 
betical sequence of the authors’ names. The botanical 
literature, on the other hand, shall simply be treated in 
chronological order, without regard to any division into periods. 
The full titles of the papers referred to are to be found on 


268 


MONTGOMERY. 


[Vol. XV. 


page 542, where their arrangement is according to the alpha¬ 
betical order of the authors’ names, both the zoological and 
botanical papers being in this one list. A certain number 
of contributions dealing with nucleoli are entered into the 
literature list, which I was unable to find in the libraries 
at my disposal; all such papers have been distinguished by 
an asterisk (*); the contents of some of the latter I have 
reviewed from the citations of other writers. 

Literature reviews are here given of all papers, with the 
object of furnishing a reference library on the subject; in 
Chapter IV, consequently, brief allusions only are made to the 
views of particular authors, and readers can compare their 
views by referring to the present section. This arrangement 
of the literature appears the most practical. 

A. Zoological Literature. 

1781-1860. 

Fontana ( 1781 , cited by Carnoy, ’ 84 ) was the first to figure 
the nucleolus in the nucleus, which he describes as “ un corps 
oviforme, pourvu d’une tache en son milieu.” 

The discoverer of the nucleolus in germinal vesicles is 
R. Wagner (’ 35 ), and he termed it “Keimfleck” or “ macula 
germinativa.” He notes that the germinal vesicle of Unio and 
Anodonta “zeigt constant zwei Flecke in Form von Kreisen, 
welche sich schneiden, selten finden sich Abweichungen; der 
grossere derselben mochte eine gewisse Aehnlichkeit mit dem 
Keimfleck haben.” In his “Nachtrag” to the same paper, he 
states: “Der Keim ist bei seinem ersten Auftreten eben das, 
was ich Keimfleck genannt habe. Es ist eine Schicht korniger 
Masse, welche bald einfach (Saugethiere, Schnecken, Insekten 
etc.) als Fleck erscheint, bald mehrere zerstreute Kiigelchen 
bildet (Flusskrebs, Fische, Batrachier), . . . die an der inneren 
Wand des Keimblaschens angeheftet sind.” In two subsequent 
communications (’ 36 , ’ 37 ) he notes the occurrence of nucleoli 
in the germinal vesicles of Coryna , Lucernaria , Cyanea , Chry- 
saora, As terms, and hisecta , and finds in Melolontha vulgaris 
one large and one small nucleolus. Finally he remarks : “ Viel- 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 269 

leicht bildet das Material des Keimblaschens und der Keim- 
flecke die Grundlage zum serosen Blatt und zum Fruchthof 
der Keimhaut.” (Jones, ’ 35 , ’ 37 , does not mention the nucle¬ 
olus;. accordingly, he is not the discoverer, as is claimed by 
Bischoff.) 

Valentin (’ 36 , cited by Carnoy, ’ 84 ) describes the nucleolus 
as a “ rundes Korperchen, welches eine Art von zweitem Nucleus 
bildet.” (On this historical ground Carnoy considers the term 
“nucleolus” should be limited to his “ nucteole-noyau.”) 

Valentin (’ 39 , mentioned by Carnoy, ’ 84 ) introduces the 
terms “ nucleolus ” and “ Kernkorperchen ” ; the latter term 
was proposed also by Schwann (’ 39 ) in the same year. 

Bischoff (’ 42 ) found in the egg of the rabbit one nucleolus, 
“ ein schwach granulirtes Kornchen,” which he considers to be 
a “ Zellenkern.” 

Vogt (’ 42 ) found several nucleoli (six to twelve) in the ova of 
Coregonus; these subsequently migrate into the yolk to form 
the first cells of the blastoderm. 

Leydig (’ 49 ) describes in the germinal vesicle of Nephelis 
one nucleolus, in Clepsine one or numerous ones, in Piscicola 
two to four, while in Haemopis “ der Keimfleck war einfach, 
8 -formig oder doppelt.” 

Kolliker (’ 49 ) studied numerous Gregarines, and concludes 
that the nucleoli (“Kornchen”) “bei manchen Gregarinen 
gewisse bestimmte Entwickelungen durchlaufen, namlich bei 
jungen Individuen einfach vorhanden sind, bei alteren allmalig 
in zwei, drei oder mehr Korner zerfalien.” In G. terebellae , 
clavata , saenuridis> and enchytraei there is a single nucleolus ; 
G. sipunculi has from one to six ; G. heeri , six to eighteen ; 
G. sieboldii , one to seven, which are either homogeneous or 
vacuolar, or else only one or two are present, and each of these 
is composed of a mass of smaller ones ; G. brevirostra has from 
six to nine nucleoli. 

Loven (’ 49 ) studied the eggs of Modiolaria , Cardmm , 
Patella , and Solen, and found that during the process of 
fecundation the nuclear membrane ruptures, and the nucleolus 
passes out through the vitelline membrane. (It is very prob¬ 
able that he confused the nucleolus with a pole body.) 


270 


MONTGOMERY. 


[Vol. XV. 


Quatrefages (’ 49 ) found that preceding the first maturation 
division of Teredo the nucleolus dissolves in the nucleus. 

v. Wittich (’ 49 ) found that in the germinal vesicles of 
Lycosa, Theridium , Epeira the “Keimfleck” first appears 
“ als ein matt gelblicher, nicht immer scharf begrenzter, aber 
durchaus homogener Fleck, wird immer entschiedener rund, 
verliert seine Homogenetat, indem er hie und da den Schein 
von unregelmassig rundlichen Aushohlungen bietet, und neben 
ihm treten zuletzt zerstreut ungleich geformte Korperchen 
auf, die dem ersteren sehr ahnlich, an Zahl immer mehr 
zunehmen, je mehr sich das Blaschen [Kern] seinem ganz- 
lichen Schwinden nahert.” In Gasterosteus aculeatus the 
number of the “ Keimflecke ” increases with the size of the 
egg. In the youngest germinal vesicles of Fringilla there is 
at first no nucleolus, later a single large, excentric one. 

Leydig (’ 50 ) finds that in the ovarial egg of Paludina vivi - 
para there are two widely separated nucleoli, while in the ripe 
egg they are in contact with each other : “ so muss wohl ange- 
nommen werden, dass der achterformige Keimfleck des reifen 
Eies durch Aneinanderriicken und theilweises Verschmelzen 
der friiher getrennten Korperchen entstanden sei.” 

Leydig (’ 52 ), ovum of Synapta digitata: there is a single 
nucleolus with a vacuole ; “ was aber als eigenthiimlich hervor- 
tritt, ist, dass er constant an einem Pol des Keimblaschens 
liegt und zwar in einer tellerformigen Grube desselben.” 

Leuckart (’ 53 ) states : “ Der Keimfleck bildet eine zusam- 
menhangende Masse von feinkorniger Beschaffenheit und 
opakem Aussehen, die unter dem Deckglaschen mancherlei 
Formen annimmt und ohne Umhiillungshaut ist. Nicht sel- 
ten lassen sich im Innern auch einzelne grossere Molekiile 
ganz deutlich unterscheiden. In manchen Fallen nehmen 
diese Molekiile an Zahl und Selbstandigkeit in einem solchen 
Grade zu, dass der ganze Keimfleck eine haufenformige 
Aggregation von Kornern darstellt.” 

Hessling (’54) finds in the youngest eggs of Unio a single 
large nucleolus ; in larger ova there is a larger and a smaller 
nucleolus, the latter having divided off from the former, and 
showing a different reaction to acetic acid. 


No. 2'.] COMPARATIVE CYTOLOGICAL STUDIES. 271 

Lacaze-Duthiers (’ 54 ) finds in the eggs of Lamellibranchs 
either one nucleolus, or when two are present they are of 
unequal size. 

Leydig (’ 55 a) says that in the egg of Cyclas “ der Keimfleck 
hat constant die Bisquitform.” In a second paper of the same 
year (’ 55 b) he makes the following notes on the ova of Rotato¬ 
ria: in Notommata myrmeleo there are about 100 finely granu¬ 
lated nucleoli ; in N. sieboldii “ Die Keimflecke erscheinen 
als Haufen von kleinen, hellen Kiigelchen,” and disappear in 
the ripe egg ; in N. centrara and Brachionus bakeri there is a 
single large nucleolus. 

Agassiz (’ 57 ) in studying the egg of the turtle introduces 
the following terms: “ ectoblast ” for cell membrane, “ meso- 
blast” for nucleus, “ entoblast,” or ‘‘Wagnerian vesicle,” for 
the nucleolus, and “ entosthoblast,” or “Valentinian vesicle,” 
for the body sometimes enclosed in the latter. In the youngest 
ova the nucleoli are absent, later they become numerous and 
large, though they disappear in the ripe egg. The excentric 
vacuole (“ Valentinian vesicle ”) of the nucleolus “ increases in 
size at a greater proportionate rate than its parent, the 
“ Wagnerian vesicle,” till at its final stage it oftentimes occupies 
three-fifths of the diameter of the generating medium.” 

Lacaze-Duthiers (’ 57 ), ovarian egg of Dentalium: at first 
there is but a single nucleolus, later a second one appears and 
apposes itself to the former ; the volumes of the two are 
different. (Cf. Fol, ’ 89 .) 

Remak (’ 58 ), blood cells of Galius: “ Es kann kaum einem 
Zweifel unterliegen, dass die Theilung der Blutzellen mit der 
Theilung des Kernkorperchens beginnt. . . . Die Regel ist, 
dass das Kernkorperchen sich in zwei Theile abschniirt, und 
ebenso der Kern in zwei Kerne. Wie es aber zuweilen vier 
Kernkorperchen giebt, so finden sich auch zuweilen vier Kerne 
in einer Zelle.” 

1861-69. 

PfUiger (’ 63 ) found one nucleolus in the egg of the calf. 
While in the “Urei” of the cat he makes the interesting 
observation that after a division of the nucleus, whereby one 


272 


MONTGOMERY. 


[Vol. XV. 


of the daughter-nuclei retained the original nucleolus, in the 
other a new nucleolus soon appeared, first in the form of a 
granular mass. 

In the paper by Balbiani (’ 64 ) movements of nucleoli are 
described for the first time, and these observations were made 
upon the living eggs. The first kinds of movements which he 
distinguishes are exhibited by the eggs of spiders : “ ces 
mouvements de la tache germinative sont caracterises par la 
production de prolongements transparents ayant presque tou- 
jours la forme de lobes arrondis qui s’allongent et se retractent 
alternativement.” The second kind of movements is shown 
in the egg of Phalangium , where there is a single large, 
spherical nucleolus, which appears spongy, owing to the pres¬ 
ence of a number of vacuoles, some of which “ s’el&vent plus 
ou moins au dessus de la surface en soulevant sous forme 
d’une ampoule la couche la plus externe de la substance du 
corpuscule. . . . Lorsqu’un porte son attention sur une de ces 
v^sicules superficielles, on ne tarde generalement pas a la voir 
grossir insensiblement, en meme temps que la couche de sub¬ 
stance qui forme sa paroi exterieure se souleve en s’amincis- 
sant de plus et plus ; puis, assez brusquement, cette paroi se 
rompt comme sous la pression d’un liquide interieur, et ses 
bords se retractent vers la base adherente de l’ampoule qui se 
trouve ainsi transformee en une petite cupule ou excavation 
superficielle, . . . et bientot il ne reste plus aucune trace de 
l’ampoule ni de F excavation qui lui a succddd.” All the periph¬ 
eral vacuoles discharge themselves thus in succession, while 
at the same time the smaller central vacuoles increase in size 
and wander towards the periphery to take the place of the 
preceding. Balbiani compares these movements to those of 
the contractile vacuoles of the Rhizopoda , but notes this dif¬ 
ference : in the latter forms the vacuole always forms itself 
at the same place again. In the eggs of Geophilus and of 
Helix pomatia he finds that the vacuole discharges through a 
small orifice. 

Balbiani (’ 65 b) describes some remarkable structures in ger¬ 
minal vesicles, all studied in life. In Geophilus longicornis 
there is an external infundibular canal extending from the sur- 


2 73 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 

i 

face of the nucleus to the surface of the vitellus, its larger 
opening being apposed to the nucleus. A smaller inner infun¬ 
dibular canal extends from the nucleolus into the outer canal. 
The numerous vacuoles of the nucleolus are contractile, and 
empty into the inner canal. He believes that these canals 
disappear at the time that the nucleolus does. “Dans les 
ovules de la Chienne, apres la separation des follicules primor- 
diaux, la vesicule et la tache germinative offrent chacune un 
prolongement canalicul^ dont Tun est int6rieur a l’autre, 
comme chez le Geophile. . . . Chez la Raie, ou les ovules 
renferment generalement d’un k quatre petits corpuscules 
germinatifs creuses d’une vacuole centrale, chacun de ceux-ci 
emet un nombre variable de petits canaux, ordinairement de 
deux a quatre, lesquels traversent dans differentes directions 
la cavite de la vesicule, percent sa paroi et vont se perdre 
dans le vitellus ambiant. . . . Chez les poissons osseux 
et les Batraciens, dont les oeufs renferment . . . un grand 
nombre de taches germinatives adherentes a la paroi interne 
de la vesicule, celle-ci est entouree d’un systeme de canaux 
rayonnants vers la surface de l’oeuf, Hg^rement flexueux et 
de longuer inegale suivant le trajet qu’ils ont k parcourir 
pour atteindre cette surface. Chaque canal est en rapport 
avec un des corpuscules precedents, et presente un calibre 
correspondant au diametre de ce dernier. . . . Quelquefois, 
ainsi que je l’ai observe chez quelques Crustac^s (Ecrevisse, 
Cancer moenas), ces taches multiples s’ont paru en outre 
reunies, dans l’intdrieur de la vesicule, par des canaux qui 
s’etendaient de l’une a l’autre. . . . Chez plusieurs Annelides, 
Turbellaries, Mollusques et Acalephes, dont j’ai examine les 
oeufs, ceux-ci ne renfermaient pour la plupart qu’une tache 
germinative simple, souvent assez volumineuse, en rapport 
avec un canal unique renferm6 dans l’interieur d’un deuxi£me 
canal emanant de la vesicule germinative.” In the germinal 
spots of Helix , Vortex , and Prostomum he noticed one or 
several contractile vacuoles. 

Schron (’65) finds in the eggs of the cat and rabbit one or 
two “ICorner” in the nucleoli of the larger eggs, though not 
in those of the smaller eggs. He considers the “Korn” dif- 


MONTGOMERY. 


[VOL. XV. 


274 

ferent in structure and substance from the rest of the nucleo¬ 
lus, and that it is characteristic for a certain stage of the cell. 

Stepanoff (’ 65 ) describes for the youngest germinal vesicles 
of Cyclas two nucleoli which are unequal in size, while in more 
mature ova there are usually two (seldom one) large ones. He 
figures, further, in one nucleus a smaller nucleolus in contact 
with a larger one. 

La Valette St. George (’66) studied in iodized serum the 
germinal vesicles of various animals. In the egg of the kitten 
there is one large nucleolus, either homogeneous or finely 
granular, containing sometimes a large vacuole. In that of 
the embryo of a sheep he noticed one or several nucleoli, with 
slight differences in size, finely granular in structure, and con¬ 
taining each a clear vacuole. In the egg of a larva of Libella 
there was a small and a large nucleolus, the latter being darker 
and more refractive, and spherical or irregular in form ; “ seine 
Substanz war entweder homogen oder zeigte je nach der Ein- 
stellung des Mikroskopes hellere oder dunklere Flecken von sehr 
verschiedener Zahl und Grosse, von unmessbarer Kleinheit bis 
zu zwei Drittel des Keimfleckes. . . . Anfangs war der grosse 
Keimfleck unregelmassig geformt fast viereckig und zeigte in 
der Mitte eine hellere Stelle, etwa ein Drittel so gross wie der 
ganze Keimfleck und daneben ein zweites kleineres Fleckchen. 
. . . Nach einer Viertelstunde hatte er seine Form geandert, 
der kleinere Fleck war verschwunden, der grossere nach der 
Spitze zu geriickt. Nach Verlauf einer halben Stunde war er 
kuglig geworden und jene helle Stelle verschwunden.” (In 
this last stage the nucleolus touches the nuclear membrane, 
according to his Fig. 2c.) In the egg of Porcellio scaber the 
nucleolus is an irregular granular mass, and later becomes a 
massive body ; “ zuweilen stellt er einen nach einer Seite 
geoffneten Ring dar, oft auch eine ausgehohlte Kugel.” By 
these observations he believes he has proved what Schron 
termed a solid granule (“ Korn ”) to be a vacuole. 

Ransom (’ 67 ), egg of Gasterosteus: young eggs with numer¬ 
ous peripheral germinal spots, which are spherical and homo¬ 
geneous. He supposes these “are soluble in some of the 
constituents of the yolk, and we may thus explain their disap- 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


2 75 


pearance in ripe ova.” A 1 . 5 ^ solution of NaCl gives rise to 
vacuoles in the nucleoli (this antedates the observation of 
Morgan, ’96). 

Van Beneden (’69) studied Greganna gigantea: “ Le nombre 
de nucldoles varie 4 chaque instant ; quelques-uns disparais- 
sent, tandis que d’autres se forment ; ils apparaissent sous 
forme d’un petit point presque imperceptible ; ce point grandit 
jusqu’a certaines limites; il devient un veritable corpuscule 
forme d’une substance homog^ne tr&s-rdfringente, puis le 
corpuscule diminue de volume ; il refracte de moins en moins 
la lumiere, enfin il disparait.” 

Clapar&de (’69) found in the egg of Lumbricns terrestris 
that the nucleolus “ ist doppelt, indem er aus zwei einander 
beriihrenden ungleich grossen Kiigelchen besteht.” 

1870-yg. 

Eimer (’71), epithelial cells of the snout of Talpa: each 
nucleolus is surrounded by a clear space (“ Hof ”), and the 
outer boundary of this space “war bezeichnet durch zahl- 
reiche kleine Piinktchen. . . . Im optischen Querschnitt 
stellten diese Kornchen einen Kreis um den hellen Hof des 
Kernes dar.” 

Eimer (’72) finds in the earlier stages of the egg of Lacerta 
that all the nucleoli are grouped near the center of the 
nucleus, while in more advanced ova there are numerous larger 
peripheral nucleoli, and smaller ones in the other portions of 
the nucleus ; around each of the large peripheral nucleoli are 
situated concentric rows of smaller ones. Here, as well as in 
Cistudo , Testudo , and Tropidonotus , the smallest nucleoli are 
homogeneous, while the larger contain vacuoles. He con¬ 
cludes that “ die complicirt gebauten Keimflecke aus einfachen 
Kornchen” are built up. 

Kleinenberg (’72) : in the egg of Hydra the single spheri¬ 
cal nucleolus contains “ein auffallend stark lichtbrechendes 
Korperchen. . . . Nach kurzer Zeit schwindet es wieder.” 
The nucleolus then becomes irregular in form, breaks into 
small granules, and he supposes that these latter become 
dissolved. 


MONTGOMERY. 


[Vol. XV. 


276 

Eimer (’73), nervous system of Beroe: each nucleus contains 
one large nucleolus. “ Aufmerksamer Beobachtung kann es 
nicht entgehen, dass jede Epithelzelle von einer Primitiv- 
fibrille versorgt wird. ... Ich kann nur so viel sagen, dass 
ich dieselbe [Primitivfibrille] stets auf das Centrum des Kerns 
zugerichtet sah, so dass ich zu der Ansicht hinneige, es werde 
sich spaterhin ihre Endigung im Kernkorperchen feststellen 
lassen.” 

Fol (’73) noticed in the egg of Geryonia fimgiformis one 
large nucleolus, containing one large, or several smaller 
vacuoles. 

Auerbach (’74). This important paper I have been unable 
to consult in the original, and quote from citations by 
R. Hertwig (’76) and Flemming (’82). According to Auerbach 
the nucleus is originally a vacuole in the protoplasm, around 
which a layer of the latter becomes differentiated to form a 
nuclear membrane. In this vacuole a nucleolus appears 
later, being derived from the protoplasm, either by a separa¬ 
tion of particles from the nuclear membrane or is produced 
out of those protoplasmic particles which had penetrated 
from the protoplasm into the original vacuole. He distin¬ 
guishes “ enucleolar,” “ uninucleolar,” and “ multinucleolar ” 
nuclei, the first being the more primitive state. The nucleo¬ 
lus has the value of an elementary organism : as long as it 
is homogeneous, it is comparable to a cytode ; when a vacu¬ 
ole appears in it, the latter stands in the same relation to 
the nucleolus as this does to the nucleus, so that that vacuole 
may be considered the nucleus (“Kern”) of the nucleolus. 
The original single nucleolus can divide into numerous nucle¬ 
oli, and the latter, by the disappearance of the nuclear 
membrane, become free, and each develops into a separate 
cell. Auerbach considers this theory as “eine vorlaufige, 
noch mit Vorbehalt aufzustellende und weiter zu priifende.” 

A. Brandt (’74) observed in life (in the blood fluid) slow 
amoeboid motions of the single nucleolus of the egg of Blatta. 

Flemming (’74) investigated the egg of Anodonta. In young 
eggs the nucleolus consists of two apposed spheres of equal 
diameter; in larger eggs one of these spheres is much larger 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 277 

than the other. “ Der kleinere Theil ist starker lichtbrechend, 
auch etwas starker tingirbar, und beim Zerdrucken resistenter 
als der grosse : beide zeigen sich hierbei als eine homogene, 
zahe Masse.” The smaller has usually one large vacuole; the 
larger has several smaller vacuoles. “ Bei Anodonta scheinen 
mir ausserhalb der Fortpflanzungszeit die beiden Theile normal 
zusammenzuhangen. . . . Kurz vor Eintritt der Befruchtungs- 
zeit gewahrt man viele (aber nur reife, grosse) Eier, an deren 
Kernkorpern eine wirkliche Trennung vorgegangen ist; aber in 
der Art, dass der kleinere Biickel stiickweise abgesprengt wird.” 

Haeckel (’74) notes in the nucleolus of some egg cells “ ein 
innerstes Piinktchen, einen Nucleolinus, welchen man Keim- 
punkt (Punctum germinativum) nennen kann. Indessen haben 
diese letzteren beiden Theile (Keimfleck und Keimpunkt), wie 
es scheint, nur eine untergeordnete Bedeutung,” only the 
yolk and the nucleus being of fundamental importance. 

Ludwig (’74) gives notes on the number of nucleoli in 
various germinal vesicles. In the Coelenterata “ Das Keim- 
blaschen umschliesst durchgangig einen einzigen Keimfleck, 
welcher haufig nochmals ein Kornchen beherbergt.” There 
is one germinal spot in Echinus , Amphidetus , So las ter, 
Branchiobdella, and in Trematodes and Rhabdocoeles. 

Van Beneden (’75) remarks in regard to the egg of the 
rabbit, that there is one nucleolus, and “deux ou trois petits 
corps arrondis qui j’ai appeles pseudonucleoles.” When the 
nucleus, during the maturation of the egg, has reached the 
“ zone pellucide ” of the yolk, “ le nucleole s’accole a la mem¬ 
brane de la vesicule du c6te de la surface de l’oeuf, \k ou la 
vdsicule est appliqu^e contre la membrane. II s’aplatit contre 
la membrane et se soude avec elle ; sa substance plastique 
s’etale en une plaque qui presente d’abord un epaississement 
median. Cette lame je Tai appetee plaque nucleolaire.” 
Shortly afterwards the latter body “ grace probablement a la 
contractilite inhdrente k sa substance, . . . se ramasse en un 
corps de forme variable, souvent ellipsoidal, quelquefois lenti- 
culaire ou en forme de calotte, que j’ai appele le corps nucl6o- 
laire.” The latter is the first pole body (“ corps directeur ”), the 
nucleoplasm plus the pseudonucleolus constituting the second. 


278 


MONTGOMERY. 


[Vol. XV. 


Eimer (’75) studied the egg cells of Silurus in eye fluid, 
and found the nucleolus to present amoeboid movements. 

Kidd (’75) found slow amoeboid movements of the nucleoli 
of the epithelial cells from the mouth of the frog. These 
cells were placed in humor aqueus, and studied on a stage 
heated to 39 0 C. 

A. Schneider (’75) says : “ Les nucleoles ne sont pas un 
element constant de la structure des Gregarines ; beaucoup 
d’especes en sont normalement privees. Dans les genres 
Clepsidrina , Enspora , Gamocystis , il n’y a jamais qu’un nucle- 
ole, permanent, tres-volumineux et spherique. . . . Dans tout 
ces genres, jamais deux individus ne sont semblables a eux- 
memes au point du nombre, de la grandeur, de la configu¬ 
ration, de Topacite ou de la transparence de leurs nucleoles.” 

F. E. Schulze (’75) noticed in life that an equal division 
of the nucleolus precedes that of the nucleus, in Amoeba 
polypodia. 

Auerbach (’76) repeats some of his previous observations 
(’74) and adds that the nucleoli show a further similarity to 
the cytoplasm, in that they have a tendency to produce 
vacuoles. 

Balbiani (’76) describes certain structures in the egg of Steno- 
bothruSy which may be chromatic filaments, though I may give 
a brief citation in regard to them in this place. The con¬ 
tents of the nucleus in the fresh state appear “rempli de 
petites hachures pales, tantot paralleles les unes aux autres, 
tantot distributes plus ou moins irregulierement dans la cavite 
nucleaire. ... A Taide de Tackle acetique, on s’assure que 
ces hachures sont determinees par les corpuscules en forme de 
batonnets etroits . . . chaque batonnet parait forme de petits 
globules reunis en strie.” At the time of nuclear division, 
these “ batonnets ” become less numerous but larger. 

Van Beneden in the same year (’76) gives the results of obser¬ 
vations on the egg of Asteracanthion. There is one large nucle¬ 
olus, and eight to fifteen small “pseudonucleoles.” He did not 
notice amoeboid motions in these, but found change of form 
and successive re- and disappearance of the nucleoli in Rana , 
PolystomuMy Gregarina, and Monocystis. “ Mais je ne doute 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


279 


pas que les differences constatees dans la forme de la tache 
germinative ne doivent etre attribues a la contractility de la 
substance des nucldoles.” The vacuoles in the nucleoli are 
probably “le rdsultat de bunion momentanee de certaines 
parties de la substance nucleolaire avec le sue nucleaire.” 
Before its disappearance the nucleolus breaks into fragments, 
which then dissolve in the “ substance nucldaire.” In this 
fragmentation one fragment is always larger than the others, 
and contains the vacuole of the primitive nucleolus; it persists 
until all the smaller fragments have disappeared. 

Biitschli (’76) found that the nucleolus disappears before the 
formation of the first pole spindle in Tylenchus, Anguillula , 
Notommata , Brachionus , Triart lira, Aphis. He mentions that 
von Siebold, in 1848 , first introduced the name “nucleolus ” for 
the micronucleus of the Infusoria , and compared it with the 
nucleoli of metazoan cells. He also cites some of the earlier 
writers who compared the pole bodies with nucleoli. 

O. Hertwig (’76) calls the nucleolus “das wichtigste Form- 
element des Kerns,” and terms its substance “ Kernsubstanz ” 
in opposition to the “ Kernsaft ” (compare his brother’s paper 
of the same year). In the process of maturation he holds that 
“der Eikern der aus dem Keimblaschen frei gewordene oder 
ausgewanderte Keimfleck ist.” He noticed vacuoles in, but not 
amoeboid movements of, the germinal spot of Toxopneustes 
lividus; he observed such motions, however, in the germinal 
spots of Rana and Pterotrachea. 

R. Hertwig (’76) terms the dense substance of the nucleolus 
“ Kernsubstanz.” “ Entweder leiten sich die vielen Kern- 
korper direkt aus dem homogenen Zustand des Kernes ab, 
indem die Aussonderung der Kernsubstanz an verschiedenen 
Punkten gleichzeitig begonnen hat ; oder die zahlreichen 
Nucleoli sind, . . . durch Theilung aus einem urspriinglich 
einfachen Nucleolus entstanden.” He believes that the 
“ Nucleoli die Trager der Kernfunction sind. . . . Somit 
miissen wir in alien den Fallen, in denen sich ein oder mehrere 
Nucleoli im Kerne differenziren, in diesen die Thatigkeitscen- 
tren des Kernes erblicken.” 

Schwalbe (’76) studied the nuclei of retinal ganglion cells of 


28 o 


MONTGOMERY. 


[Vol. XV. 


the ox, rabbit, and sheep: in the smallest nuclei there is no 
nucleolus within the nucleus, but there are small peripheral 
prominences on the inner surface of the nuclear membrane; 
when a nucleolus is present within the nucleus it is jagged in 
outline, with fine, thread-like processes. The substance of the 
nuclear membrane “ stimmt in alien Eigenschaften mit der des 
Kernkorperchens vollstandig iiberein, und ist mit ihr continu- 
irlich.” Further, the substance of the peripheral prominences 
is quite identical with that of the nucleolus, and “ Man konnte 
in dem Falle, wo ein innerer Nucleolus fehlt, geradezu davon 
reden, dass als Ersatz dafur wandstandige Kernkorperchen vor- 
handen seien.” In similar cells of the calf, there are no nucleoli 
in the smallest nuclei; in larger ones there are from two to 
four, one or two lying within the nucleus, the others being mere 
thickenings of its membrane; “beim Wachsen des Kernes 
(12.5/4) nimmt die Hohe und Zahl dieser Wandverdickungen 
immer mehr ab, wahrend im Innern ein gut ausgebildeter 
zackiger oder eckiger Nucleolus von 2.7 bis 3.6/4 das gewohn- 
liche ist/' He considers the substance of the nucleoli and of 
the nuclear membrane to be at first identical, and to be diffused 
in the “ Kernsaft.” In the sympathetic ganglion cells of the 
frog, he noticed, on the heated stage, that the nucleoli exhibited 
slow changes of form; and in these nuclei he distinguishes 
“ Nucleolarsubstanz, den Kernsaft und die reticulare Sub- 
stanz.” 

O. Hertwig (’77a) found in the egg of Haemopis one large 
nucleolus, with usually one large vacuole; and also a number of 
small nucleoli, some of which contain each a small central 
vacuole. In the production of the pole bodies: “Aus den 
Theilstiicken des Nucleolus und einem Rest des Kernsaftes 
entsteht ein fasriger, spindelformiger Kern . . . es muss dahin- 
gestellt bleiben, ob der ganze Nucleolus oder nur ein Theil des- 
selben und ob die Nebenkiigelchen [Nebennucleolen ?] in die 
Zusammensetzung der Spindel mit eingehen.” 

v. Kennel (’77) remarks of the ripe egg of Malacobdella: 
“ der Kern . . . enthalt eine mehr oder minder grosse Anzahl 
stark lichtbrechender runder Tropfchen, die sich meist an 
seiner Peripherie befinden.” 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 281 

Mark (’ 77 ) : the salivary gland cells of Chionaspis contain 
each forty to fifty nucleoli; corresponding cells of Aspidiotus 
have a single large one which may contain from two to seven 
“ nucleoluli.” In cells of the oval gland of Chionaspis the 
nucleus contains a true nucleolus, usually without nucleoluli, 
and also a “ Fetttropfchen,” which differs from the former in 
color and refraction. (The Fig. 32 of the salivary gland cells 
of AspidioUis shows each nucleus to contain a double nucleolus, 
containing a larger sphere apposed to, in one case separate 
from, a smaller colorless sphere.) 

A. Brandt (’ 78 ) gives observations on the germinal vesicles 
of different forms. In Aeschna grandis : “Der vom Keimblas- 
chen umschlossene Keimfleck ist wie dieses, urspriinglich 
rund, aber in noch viel hoherem Grade, und zwar unstreitig 
bei alien von mir beobachteten Insekten, amoboid beweglich, 
so dass seine Form meist sehr verschieden erscheint. Nicht 
selten ist er in einige Theile zerfallen. ... In einzelnen 
Keimblaschen lagen ausser dem Keimflecke noch ein oder 
mehrere Kornchen von verschiedener Grosse ; — nur ein Paar 
Keimflecke wurden aufgefunden, welche anscheinend aus zwei 
aneinandergedrangten und theils ubereinander geschobenen 
Kugeln bestanden.” In Periplaneta vacuoles as well as solid 
“ secundare Keimflecke ” occur in the nucleolus. In the egg 
of Nemura, after the action of acetic acid, the vacuoles in the 
nucleolus increase in size and in each a small granule is to be 
seen. In Gryllus , Lepisma , and Holostomis the germinal spot 
is amoeboid : “ Die amoboide Beweglichkeit veranlasst nicht 
selten das Loslosen einzelner Partikel, welche, wie der Keim¬ 
fleck selbst, amoboid-contractil sind. Die Zahl und Grosse 
dieser gelegentlich wieder zusammenfliessenden Partikel ist 
eine ausserst verschiedene thus the nucleolus may break 
into a number of equal-sized pieces, or into a mass of very 
fine granules. In the egg of Tegenaria there is usually a 
single vacuolated nucleolus, though sometimes there may 
be present also two “ Nebenkeimflecke.” In Distonmm the 
«Keimfleck . . .ist in sehr hohem Grade mit amobenartiger 
Beweglichkeit begabt,” and there is a central body in the 
nucleolus which changes its form periodically. Brandt observes 


282 


MONTGOMERY. 


[Vol. XV. 


in regard to the frog’s egg : “Der Keimfleck des Froscheies, 
in den allerjiingsten Eianlagen meist ein zusammenhangendes 
Gebilde, erscheint bekanntlich spater, in eine grossere Anzahl 
von rundlichen Klumpchen zerfallen — und diese fand ich 
(bei Rana esculenta) amoboid gestaltet”; and adds, against 
Biitschli (’76), “ist ein£uwenden, dass dieser Zerfall des Keim- 
flecks als amoboide Erscheinung keineswegs auf ein Absterben, 
sondern im Gegentheil auf eine erhohte Lebensthatigskeit 
hinweist.” 

Brock (’78) : the immature ovum of Anguilla has one or two 
large nucleoli ; the number of the latter increases with the 
size of the egg. 

Eimer (’78) notes the great relative and absolute size of the 
nucleus and nucleolus in ganglion cells, and finds it to be 
paralleled only in egg cells. 

O. Hertwig (’77b, ’78a) noticed in the nucleolus of the 
maturing egg of Asteracanthion certain changes, “die darin 
bestehen, dass die in seinem Innern bisher zahlreich vorhan- 
denen kleinen Vacuolen verschwinden und in seiner Mitte oder 
mehr der Peripherie genahert eine grossere Vacuole erscheint, 
die fast ganz von einem kugligen aus Kernsubstanz bestehenden 
Korper erfiillt wird. . . . Plotzlich verschwinden die in ihm 
gelegenen Vacuole mit ihrem kugligen Korper unter dem Auge 
des Beobachters,” and in consequence the nucleolus begins to 
gradually shrink in size, and hours afterwards has com¬ 
pletely disappeared. The body within the large vacuole of the 
nucleolus corresponds to the smaller, more deeply staining 
portion of the original nucleolus, and during the nuclear division 
reaches out of an opening in the vacuole beyond the surface of 
the nucleolus, takes on the form of a long, thin rod, and occupies 
the middle point of the first pole spindle; while at the same 
time the remaining portion' of the nucleolus gradually breaks 
into a granular mass, which then disappears. Also in Sphae- 
rechinus , Ascidia, some Coelenterata , and various Mollusca , he 
noticed a similar differentiation of the nucleolus into two sub¬ 
stances, namely, a smaller, deeply staining portion apposed to, 
or enclosed by, a lighter, larger portion. 

O. Hertwig, in still another paper (’78b), investigated the 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 283 

germinal vesicles of various animals. In Eucope poly sty la there 
is one nucleolus in small eggs, several in riper ones: “ Es 
Hess sich hier feststellen, dass die zahlreichen Nucleoli durch 
Abldsung vom urspriinglichen einfachen Keimfleck entstehen.” 

Klein (’78) studied the stomach cells of the newt, and con¬ 
cludes “that in most cells the so-called nucleoli are local 
accumulations of the intranuclear network, that they are incon¬ 
stant in size and number, and that they are only transitory 
appearances.” 

Schindler (’78), Malpighian tubules of insects : after a cell 
has become obliterated by the outflow of its secretion, its 
nucleus becomes a new cell, and its nucleolus a new nucleus. 

Whitman (’78) found in the egg of Clepsine one to three 
nucleoli, each “ composed of several highly refractive pieces.” 

Bergh (’79) found in the egg of Gonothyraea (Campanularia ) 
a single large nucleolus, which is usually round, but sometimes 
with irregular outlines caused by slow amoeboid movements 
(observed in life), these motions being most vigorous later, 
when the nucleolus begins to divide. It increases in size, and 
acquires one or two vacuoles. In a later stage, but before the 
production of the pole bodies, there are a number of irregular 
nuclear bodies (staining as the original nucleolus), which had 
been produced by division of the nucleolus; in one case he 
actually observed the division of the nucleolus, which lasted 
half an hour, and at the same time the vacuole of the primitive 
nucleolus seemed to divide into two, so that each daughter- 
nucleolus received a daughter-vacuole. “ Oft macht es den 
Eindruck, als ob das Volum der secundaren Keimflecke zusam- 
mengenommen grosser ware, als das der primaren fur sich 
. . . eine active Wanderung der Nucleoli durch den Kernsaft, 
wie dies Auerbach [’74] bei gewissen Nematoden in den Vor- 
kernen gesehen hat, kommt wahrscheinlich hier nicht vor.” 
The nucleolus also divides in the egg of Clava. In the eggs 
of Psammechinus and Echinocardium , the single nucleolus 
begins to fragment before the chromatic network has disap¬ 
peared. The Phallnsia egg contains one large germinal 
spot, which probably disappears without fragmenting: “ich 
habe namlich unter Eiern, die im Keimblaschen einen scharf 


284 


MONTGOMERY. 


[Vol. XV. 


begrenzten, durch die Osmium-Carminbehandlung rubinroth 
gefarbten Keimfleck zeigten, auch solche gefunden, welche 
statt dessen eine sehr feinkornige, bisweilen rubinroth, bisweilen 
weniger intensiv rothgefarbte Masse enthielten, die nicht scharf 
contourirt war, aber von derselben Grosse wie der Keimfleck. 
Falls diese Deutung, es schwinde der Keimfleck ohne sich 
vorher zu theilen, richtig ist, beginnt die Auflosung desselben 
mit dem Schwinden der Vacuolen in seinem Innern.” 

Klein maintains his previous views in regard to the nature 
of the nucleoli in two papers published in the following year 
(’79a, ’79b). 

1880 . 

Van Beneden (’80) studied the egg of the bat, and found one 
nucleolus (rarely two): “ on trouve en outre quelques granules 
tres petits, tous d’egales dimensions, repandus dans le corps 
de la vesicule (pseudonucldoles) ”; the latter have no resem¬ 
blance to any part of the chromatic filament. 

Biitschli (’80) incorporates in his great “ Protozoenwerk ” the 
observations of preceding authors. In Hyalosphenia there 
may be as many as six spherical nucleoli; in certain other 
Rhizopoda the “ Binnenkorper kann den von der Kernhiille 
umschlossenen Raum nahezu vollig ausfiillen.” In the Hell - 
ozoa the nucleoli are much as in the preceding group. In the 
Radiolaria (for which Biitschli follows some of the observa¬ 
tions of R. Hertwig, ’79) there is usually a number of rather 
large nucleoli, frequently containing vacuoles. The nucleus 
of Thalassicola “ enthalt einen ansehnlichen, strangformigen 
und unregelmassig verastelten Nucleolus, dessen Masse nicht 
ganz homogen, sondern ausserlich feinkornig ist”; it later 
breaks into a number of segments. In Acanthometra the nucleo¬ 
lus is at first spherical, while later “ Aus dem Nucleolus-Pol, 
welcher der Einstiilpungsstelle der Kernmembran zugewendet 
ist, bildet sich eine helle homogene Masse aus, welche den 
dunkleren Haupttheil des Nucleolus wie eine Kappe bedeckt 
oder auch wie eine Vertiefung desselben eingesenkt erscheint. 
Der Nucleolus erscheint demnach jetzt von zwei verschiedenen 
Substanzen zusammengesetzt.” In many Flagellata a nucleo- 


No. 2 .] COMPARATIVE CYTOLOGICAL STUDIES. 285 

lus is absent, in others there is a single one, sometimes with 
a vacuole; in the Choanojiagellata there is always one large, 
spherical nucleolus, in the Cystojlagellata several of various 
sizes; and in the Dinojiagellata there may be several small 
nucleoli, which are sharply localized from the chromatin, but 
show the fine reticulation of the latter element. In the Ciliata 
and Suctoria there are nucleoli of varying size and number in 
the macronucleus, but none in the micronucleus. 

Chun (’ 80 ) finds in the egg of all Cte7iophora a single large 
nucleolus, very rarely two. 

Engelmann (’ 80 ) figures the nucleoli of certain ciliated cells 
of various invertebrates as each surrounded by a clear space, 
the outer boundary of which is marked on optical cross-section 
by a circle of granules. 

Flemming (’80) concludes in regard to the nature of the 
nucleolus: “ Dass die Nucleolen iiberhaupt keinerlei mor- 

phologischen Antheil an der Kernvermehrung nehmen”; and 
“ Dass die Dinge, die wir Nucleolen nennen, vielleicht gar keine 
morphologisch wichtige Theile des Kerns, sein mogen, sondern 
nur Ablagerungen von Substanzen, welche fur den Stoffwechsel 
im Kern verbraucht und wieder neugebildet werden; sie wiirden 
damit gewiss physiologisch wichtige Theile des Kerns bleiben, 
— was ohnehin durch ihr fast allgemeines Vorkommen bewahrt 
wird, — aber doch keine eigentlich organischen, d. h. morpholo- 
gisch-wesentlichen Kernbestandtheile.” 

O. Hertwig (’ 80 ) found in the eggs of Chaetognatha numerous 
small nucleoli. 

Shafer (’ 80 ), ovum of Gallns: there is a single nucleolus, 
which in young germinal vesicles consists of a homogeneous 
matrix which stains slightly with haematoxylin, and a number 
of coarse granules which stain deeply; in larger ova the 
nucleolus is homogeneous throughout and stains deeply. The 
threads radiating from the periphery of the nucleolus may be 
either artefacts or may be regarded as extrusions of the homo¬ 
geneous substance of the nucleolus. Ovum of Lepus: in 
younger nuclei the nucleolus has the same general structure 
as in the fowl, though it is more irregular in form. In some 
larger ova the nucleolus “ is represented by a number (a dozen 


286 


MONTGOMERY. 


[VOL. XV. 


or so) of globules of varying size which appear to lie loose 
within the germinal vesicle. An intravesicular network is 
sometimes present, and serves to unite the granules of the 
macula. ... It is possible that the homogeneous matrix 
above described may represent the remains of such a network, 
the filaments of which have shrunk up into a mass on contact 
with the hardening reagent ” (picric acid and alcohol). 

Trinchese (’ 80 , according to Platner, ’86) found in the germinal 
vesicle of Amphorina coerulea a “macchia germinativa laterale 
o accessorial and a “macchia germinativa principale,” the 
latter being about seven times the size of the former. 

1881 . 

Balbiani (’ 81 ) investigated the salivary gland cells of the 
Chironomus larva. There are here “ Deux gros nucleoles irre- 
guliers, larges de 0.03 a 0.04 mm., bosseles a leur surface, et 
formes d’une substance refringente granuleuse, creusee d’un 
plus ou moins grand nombre de vacuoles isolees ou confluentes. 
II arrive assez souvent que les deux nucleoles se confondent par 
une partie plus etroite qui les reunit comme une sorte de pont; 
d’autres fois enfin, ils se fusionnent plus ou moins intimement 
en un seul nucleole, dont le diametre est le double de celui des 
nucldoles isoles.” The ends of the chromatin filament are 
apposed against the nucleolus; and the latter differs both 
chemically and morphologically from this “ cordon nucleaire.” 

Giard (’ 81 ) observed in the egg of a Spionid during life a 
single central nucleolus. A certain time before completed 
maturation an “Element cellulaire ” appears in the nucleus, 
which is a little smaller than the latter, and encloses in its 
center a small “ noyau ” : “ D’abord fort eloigne du nucleole, il 
s’en approche progressivement et vient s’appliquer a sa surface, 
ou il s’aplatit et prend la forme d’une double calotte. En 
s’appliquant de plus en plus contre le nucleole, il perd son 
noyau et finit par se reduire a une double membrane qui entoure 
le nucleole,” . . . and finally its substance fuses with that of 
the nucleolus. 

Hubrecht (’ 81 ), egg of Proneomenia : “ in all the different 
stages of development of the ovum the germinal spot is double: 


No. 2 .] COMPARATIVE CYTOLOGICAL STUDIES. 287 

a larger and a smaller sphere may be distinguished, which, 
however, are not connected in any way whatever . . . but 
perfectly free and independent of each other.” 

Mark (’81) finds that during the maturation of the egg of 
Limax campestris the male as well as the female pronucleus 
may contain as many as fifty or sixty “pronucleoli,” which dis¬ 
appear before the copulation of the two pronuclei. In an 
undetermined species of Limax he “ observed in both female 
and male pronuclei a single nucleolus of much greater size and 
more deeply stained than the other nucleoli.” 

Pfitzner (’81) finds that the “ Kernsubstanz ” is contained in 
the reticulum and the nucleoli; the latter lie within the meshes 
of the former, and their role is problematical. “ Wahrend des 
weiteren Verlaufes der Karyokinese verschwinden sie, werden 
anscheinend allmalig aufgezehrt, ohne direkt mit dem Geriist 
in Verbindung getreten zu sein.” 

Retzius (’81, cited by Van Bambeke, ’85): the nucleoli are 
simple local accumulations of the chromatin, derived from the 
nuclear reticulum. 


1882 . 

Blochmann (’82) observed in the egg of Neritina one large 
nucleolus containing a vacuole. Preceding the pole body pro¬ 
duction, the nuclear membrane vanishes, and the nucleolus at 
first retains its original size, then breaks up into several equal¬ 
sized fragments. “ Dass die Elemente der Kernplatte aus 
Theilstiicken des Nucleolus entstehen, kann bei unserem 
Objekt keinem Zweifel unterliegen, da ich alle Uebergangszu- 
stande vom unversehrten Nucleolus bis zur ausgebildeten 
Kernplatte beobachtet habe.” After the two pole bodies have 
divided off, the remaining chromosomes in the female pro¬ 
nucleus fuse together to form a deeply staining, spherical 
body, which resembles the original nucleolus. 

Flemming in his classical work (’82) gives the following defini¬ 
tion of nucleoli: “ Substanzportionen im Kern von besonderer 
Beschaffenheit gegeniiber dem Geriist und dem Kernsaft, fast 
immer vom starkeren Lichtbrechungsvermogen als beide, mit 


288 


MONTGOMERY. 


[Vol. XV. 


glatter Flache in ihrem Umfang abgesetzt, stets von abgerun- 
deter Oberflachenform, meist in den Gerustbalken suspendirt, 
in manchen Fallen ausserhalb desselben gelagert.” A mem¬ 
brane is absent around all nucleoli. He (erroneously) attrib¬ 
utes the discovery of the nucleolus to the botanist Schleiden. 
Flemming holds it probable that with the possible exception of 
spermatozoa one or more nucleoli occur in every nucleus, of 
which it is therefore an important organ (in this conclusion 
he departs from the views expressed in his previous contribu¬ 
tion, ’80). “Die Zahl ist bei Thierzellen selten iiber 8 (mit 
Ausnahme der Kerne meroblastischer Eier), bei den meisten 
Arten von Thierzellen durchschnittlich 3 - 5 . ... Es ist der 
haufigste Fall, dass einer der Nucleolen an Grosse besonders 
vorwiegt,” this being then the “ Hauptnucleolus,” the others 
“ Nebennucleoli.” In the “ Hauptnucleolus ” of the egg of 
Lepus two parts are distinguishable, but he leaves it undecided 
whether “die Unterscheidung von Haupt-und Nebennucleolen 
eine durchgehende Geltung beanspruchen kann.” This inves¬ 
tigator notes further: “Die absolute Grosse der Nucleolen 
steht bei den meisten Zellenarten in annahernder Proportion 
zur Grosse der Kerne selbst.” The nucleolar vacuoles are 
filled with fluid. In regard to the apparent clear spaces around 
nucleoli, we read “ dass dieses Phanomen nichts anderes ist als 
ein Reflex, bedingt durch die rundliche Flache und starkere 
Lichtbrechung des Nucleolus.” He did not find amoeboid 
changes of form, but concedes that they may occur. The true 
nucleolar substance differs from the chromatin. The nucleoli 
are “ specifische Produkte des Kernstoffwechsels und zugleich 
auch specifische Formtheile des Kerns ... so kann man die 
Nucleolen ganz wohl Organe des Kerns oder der Zelle nennen.” 
They appear to be “ besondere Reproductions- und Ansamm- 
lungsstellen des Chromatins. . . . Entweder ist also in den 
Nucleolen noch ein anderweitiges Substrat vorhanden, in 
welchem das Chromatin verarbeitet wird und mit dem es in 
ihren durchlagert liegt, oder ... die Substanz der Nucleolen 
mag zwar in sich homogen sein, ist aber dann nicht identisch 
mit Chromatin resp. Nuclein, sondern eine chemische Modifi¬ 
cation, Vorstufe oder Doppelverbindung derselben.” 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 289 


Graff (’82) figures in the eggs of Proporus , Plagiostoma , and 
Vorticeros a single nucleolus containing vacuoles. 

Nussbaum (’82) studied the nuclei of gland cells (stomach 
nucosa of various Vertebrate epidermis glands of Argulus ). 
“ Es Hess sich im Allgemeinen feststellen, dass wahrend des 
ungestorten Ablaufs der Secretion die mononucleolaren Kerne 
vorherrschten, dass nach langerem Hunger die multinucleo- 
laren Kerne an Zahl vermehrt waren. . . . Ein Driisenzellenpaar 
der Saugscheibe von Argulus foliaceus hatte am 12 . Oktober 
mononucleolare Kerne; am 18 . Oktober zeigten sich viele 
Kernkorperchen.im Kern ; nach und nach ging die Granulierung 
der Zellen, die Strahlung verloren und die Kerne waren platte 
Ovoide mit mehreren glanzlosen Korperchen darin.” From 
these observations Nussbaum concludes: “ So wird man den 
Kern mit vielen Kernkorperchen als den Ausdruck einer Ruhe- 
pause der Kernfunctionen auffassen konnen, die entweder zum 
kraftigen Leben oder zum Tode iiberleitet.” 

Rauber (’82) figures the nucleoli in the ova of various verte¬ 
brates, and distinguishes the following kinds of nuclei, with 
regard to the mode of distribution of the “ chromophile Sub- 
stanz” (chromatin together with pyrenin): “globulare,” “tra- 
bekulare, “ filofde,” and “gemischte.” 

Seeliger (’82) finds that in Clavelina the nucleus of the loose 
mesoderm cell (from which the ovum is derived) becomes the 
nucleolus of the ovum, and its cytoplasm becomes its nucleus. 
In the germinal vesicle there is then one large nucleolus, in 
which nucleolini lie, and also (to judge from his figures) 
vacuoles. 

Vejdovsk^ (’82), egg cells of Sternaspis scutata: the young 
nucleus contains at first one small nucleolus, bounded by a 
membrane (though the latter structure would appear from his 
figures to be a clear space enveloping the nucleolus). “ Beim 
fortschreitenden Wachstum des Keimblaschens vergrossert 
sich auch der Keimfleck, und zwar in der Weise, dass die ihn 
umgebende Membrane einseitig sich verdickt und schliesslich 
auf dem runden sich in Pikrokarmin stark farbenden Keimfleck 
als ein glanzendes, gelbliches Buckelchen erscheint.” The 
nucleolus disappears in the ripe egg. 


290 


MONTGOMERY. 


[VOL. XV. 


1883 . 

Balbiani (’83) renewed his observations on the egg of Geophilus 
longicornis , making several emendations. In very young eggs 
there are two or numerous nucleoli, in larger eggs only one large 
one, containing one or several vacuoles. In his previous paper 
referred to, he assumed that the double tubular structure in 
these eggs served for the purpose of an intraovular circulation; 
but in the present paper he offers another explanation: that 
the double tubular structure later develops into a knotted cord, 
the distal portion of which then divides into irregular frag¬ 
ments, which become scattered through the yolk ; and then each 
of these fragments, with the exception of one which becomes 
the “ noyau vitellin,” differentiates into cytoplasm, nuclear and 
nucleolar substance, and then represents a cell of the follicular 
epithelium. 

Van Bemmelin (’83) states of the eggs of Brachiopoda : “ Sie 
haben meist zwei Kernkorperchen, die enganliegend und stark 
lichtbrechend sind. Ausser diesen nimmt man oft noch mehrere 
lichtbrechende Kiigelchen in dem gefarbten Inhalte der Eikerne 
wahr. Von Boraxkarmin werden sowohl diese Korperchen als 
die Nucleoli stark gefarbt.” (Certain of his figures show one 
of the nucleoli imbedded in another.) 

Van Beneden (’83), ovum of Ascaris megalocephala: there is 
a single “ corpuscle germinatif,” which contains all the chro¬ 
matin of the nucleus, and is contained within a special portion 
of the nucleus termed the “ prothyalosome ”; from one to three 
“ pseudonucleoles” also occur in the nucleus, but they play no 
important part in the maturation of the egg. 

Fol (’83a), egg of Ciona intestinalis: there is here one large, 
very refractive nucleolus containing a number of vacuoles 
which he believes are artefacts, since they cannot be found in 
the living egg, though their appearance after the action of 
reagents would show that the substance of the nucleolus is 
chemically not homogeneous. The nucleolus consists of a more 
refractive cortical substance, and of a less refractive, clearer 
medullary portion ; in the latter, the vacuoles are produced. 
Fol maintains that the follicle cells arise by budding from the 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 291 

egg nucleus : “ Ce nucleole a une tendance bien evidente a se 
placer dans le voisinage immediat des noyaux folliculaires en 
voie de formation. Le fait n’est pas constant, mais il est trop 
frequent”; he did not actually observe that the nucleolus gives 
off a part of its substance to the follicle cell, but supposes this 
to be the case. 

Fol, in a second paper (’ 83 b) of the same year, finds that in 
Ciona during the “production endogene” of the follicular cells 
a segment (diverticulum) of the egg nucleus breaks off, while 
the (then peripherally situated) nucleolus gives a part of its 
substance into this diverticulum, and the nucleolus then wanders 
back to another portion of the nucleus. “Chez Ascidia mam - 
viillata , le bourgeonnement de l’enveloppe a lieu simultanement 
en une foule de points, et il est tout ou moins admissible que la 
substance de la tache germinative dispersee a la formation de 
ces bourgeons.” 

Gruber (’ 83 ) describes in Actinosphaerium the growth of a 
supposed nucleolus and its division during mitosis into two 
equatorial plates; though his figures would show that he 
mistook true chromatin masses for a nucleolus. 

Jensen (’ 83 ) studied the ovum of Cucumaria; there are from 
fifteen to thirty nucleoli flattened against the nuclear membrane, 
and containing vacuoles. As shown by treatment with acetic 
or picrosulphuric acid, the outer layer of the nucleolus seems to 
be a continuation of the nuclear membrane, so that the inner, 
less refractive portion of the nucleolus appears to be situated in 
a depression of the outer surface of the nuclear membrane. 

La Valette St. George (’ 83 , quoted after Platner, ’86) found in 
the egg of an Isopod one nucleolus which is at first homogene¬ 
ous, later granular, and which may enclose a vacuole and show 
amoeboid movements. In other cases there are either several 
smaller vacuoles or one or two larger ones. 

Leydig (’ 83 ), from comparative studies, concludes that the 
nucleoli “sind Theile des Kernnetzes,” and that each of them 
is enclosed in a small, clear cavity of the nucleus. “ Die Nucle¬ 
oli konnen als eine Vielzahl von Kornchen erscheinen, die 
unter sich gleichwerthig sind. . . . Nicht selten lasst sich bei 
genauem Zusehen in der Menge kleiner und unter sich gleicher 


292 


MONTGOMERY. 


[Vol. XV. 


Kernkorper ein grosserer Nucleolus . . . auffinden (Epithel des 
Eierstocks von Aglia tau ). . . . Wahrhaft riesige Kernkorper 
kommen zu Stande, wenn viele Nucleoli zu einem einzigen 
Korper zusammenfliessen. . . . Priifen wir Herkommen und 
Beschaffenheit der Kernkorper naher, so ist beziiglich der 
kleineren Nucleoli leicht festzustellen, dass sie aus Verdich- 
tungen oder Knotenpunkten des Kernfadennetzes den Ursprung 
nehmen. Daher schon im frischen Zustande solche Kernkor- 
perchen einen zackigen Saum haben, auch durch Spitzen und 
Striche sich verbinden, die bis zum Rande des Kernes gehen. 
Aber selbst die grosseren Nucleoli . . . erweisen sich als Um- 
bildungen von Partien der Kernfaden.” In the ganglion cells 
of the brain of Limax and Avion the nucleoli are jagged in out¬ 
line, with long fibers. In the cells of the salivary gland of Nepa 
they are three or four in number, bent and elongated in form. 
Those of the corresponding gland in Naucoris have often the 
shape of a half ring, or may be lobular or band shaped, with 
cross striation. In the salivary gland of Chironomus plumosus 
there is usually a single nucleolus, spherical, lobular or tubular, 
its radiating cavity filled with a homogeneous, refractive sub¬ 
stance ; its wall contains vacuoles, “ und starke Linsen lassen 
deutlich werden, dass der ganze Kernkorper eben wieder 
die Struktur eines Schwammgebildes besitzt.” Besides the 
nucleoli there are in these cells several looped or contorted 
bodies, one of which is always in connection with the nucleolus, 
and all of which evince a cross striation, the nature of which is 
as follows : “ Mit Tauchlinsen unterscheiden wir abwechselnd 
je eine dunkle und helle Querlinie und sehen die erstere, welche 
leicht gekerbt ist, zusammengesetzt aus einzelnen kleinen 
Stuckchen, vergleichbar den Elementen einer Muskelscheibe. 
Die feinen Abtheilungslinien der den Querstrich bildenden 
Stuckchen erstrecken sich ferner durch die helle Zwischenzone, 
so dass dadurch auch eine Art von zartesten Langslinien zum 
Ausdruck kommen kann.” He believes that these cross-stri¬ 
ated structures “durch Umbildung des den Kern durchziehen- 
den Maschenwerkes entstanden sind.” In young larvae these 
structures are not seen immediately in life, but “nach und 
nach, wahrend das Thier noch lebt, tauchen die querstreifigen 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


2 93 


Bildungen auf. . . . Man darf wohl annehmen, dass die frag- 
lichen Gebilde, bevor sie dem Auge sichtbar werden, schon 
dagewesen sind und nur erst jetzt sich abheben, weil die Licht- 
brechungsverhaltnisse sich geandert haben.” Similar cross- 
striated bodies were noticed in the cells of the Malpighian ves¬ 
sels of Chironomus. In the ovarial egg of Libella Ley dig 
found one nucleolus, which consisted of a mass of granules 
grouped around a central cavity, these granules being connected 
together by fine threads ; “der lebende Nucleolus zeigt ferner 
langsam ablaufende Gestaltsveranderungen, wobei sich nach 
und nach einzelne Kliimpchen mehr oder weniger absondern.” 

Ogata (’ 83 ) investigated the pancreas cells of man, which had 
been treated with various poisons and with the induction current, 
then fixed in aqueous solution of corrosive sublimate, and with 
osmic acid. One to more than eight nucleoli may be present: 
“ Die einen farben sich wie die Kernmembrane tief mit Haema- 
toxylin. . . . Die anderen oder vielmehr das andere, denn es ist 
in der Regel nur eins, farbt sich nicht mit Haematoxylin, son- 
dern mit Eosin. . . . Manchmal hat es einen ganz feinen blauen 
Saum, als habe es selbst wieder eine Membran. Es ist viel gros¬ 
ser als die anderen Kernkorperchen, und das Feld, in dem es 
liegt, ist durch eine starkere Linie von dem ubrigen Kern 
getrennt. . . . Man wird es am unbefangensten wegen seiner 
Farbung als Plasmosoma von den ubrigen die Kernfarbung 
annehmenden Karyosomen des Kerns unterscheiden.” Some¬ 
times several smaller plasmosomata are also present. Close to 
the nucleus is a body he terms “Nebenkern,” which stains as 
the plasmosoma, but is much larger, and is apposed to the sur¬ 
face of the nucleus like a hat ; its substance is homogeneous, 
refractive, enclosing small cavities in which minute spherules 
occur, the latter having a resemblance to zymogen granules. 
The “Nebenkern” is produced by a plasmosoma which has 
wandered out of the nucleus, and there becomes the nucleus of 
a new cell. (This process is called “Zellneuerung”) 

Pfitzner (’ 83 ) found in the resting nuclei of the ectodermal 
cells of Hydra usually one central, spherical nucleolus. Its sub¬ 
stance is not identical with the chromatin in the resting stage 
of the nucleus, but becomes metamorphosed into the latter 


294 


MONTGOMERY. 


[Vol. XV. 


substance during the following mitosis ; wherefore he suggests 
the term “ Prochromatin ” for nucleolar substance. In the pro¬ 
phase of the mitosis only one nucleolus is present in the 
nucleus, while in the “ Riickkehr der Tochterkerne zum Ruhe- 
stadium waren dagegen stets mehrere vorhanden. In einem 
gewissen Stadium, wo die Nucleolenbildung beginnt, ist eine 
ganze Anzahl vorhanden ; jemehr sich der Tochterkern dem 
Ruhestadium nahert, desto mehr vermindert sich die Zahl 
unter gleichzeitiger bedeutender Grossenzunahme der iibrigge- 
bliebenen, bis fur das ausgesprochenste Ruhestadium das Vor- 
handensein eines einzigen grossen central gelegenen Nucleolus 
geradezu typisch wird,” and this he concludes to be a process 
of fusion. The nucleolus plays only a passive role in mitosis, 
“namlich die eines aufgespeicherten Nahrungsmaterials zur 
Neubildung von Chromatin.” 

Rein (’ 83 ) studied the eggs of Lepus and Cavia. In each 
there is one large nucleolus which disappears during matura¬ 
tion and is succeeded by several smaller ones, which have the 
same consistency as the first, and at the time of their first 
appearance occupy a central position in the nucleus. “ So weit 
ich den Vorgang am Saugethiere verfolgen konnte, machte mir 
derselbe eher den Eindruck eines successiven Zerfalles des 
urspriinglichen Keimflecks in immer kleinere Stiickchen, welche 
schliesslich in der Substanz des Keimblaschens verschwinden.” 

Route’s (’ 83 ) conclusions are, in the main, confirmatory of 
Fol’s (’ 83 ) observations in regard to endogenous cell formation. 
In the egg of Ciona there is one large and two or three smaller 
nucleoli, the latter being “ formes pendant revolution des 
cellules endotheliales en cellules ovulaires.” In eggs a little 
larger these “nucleoles adventifs ” become more numerous (five 
to six), and certain of them show a limiting membrane. Later 
still some of these adventive nucleoles are found in the yolk, 
where each becomes surrounded by a clear zone; these he con¬ 
siders at this stage to be the nuclei of endogenetically formed 
cells (follicular cells), the clear zone around each representing 
its cytoplasm. 

Schauinsland (’ 83 ) noticed in the egg of Distomum a single 
large nucleolus. 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


295 


A. Schneider (’83) studied Klossia, one of the Coccidia. One 
or several nucleoli are present, “ formant un ensemble souvent 
tres complexe que j’appelerai le corps nuctiolaire” Sometimes 
the largest nucleolus is enveloped on one side by a number of 
secondary, much smaller ones (“nucldolites ”), which are portions 
loosened from the inner substance of the large nucleolus, from 
which they break out through a “ canal micropylaire ” (such a 
canal was not observed in life, and on only a single fixed prepa¬ 
ration; cf. his Fig. 7 ). “ Correlativement a la multiplication du 
corps nucleolaire, le nucleole principal diminue de volume. . . . 
Touts les petits nucleoles qu’on observe dans le corps nucleo¬ 
laire me paraissent descendre aussi surement du nucleole 
primitif ou ancetre que les jeunes d’une espece de leurs parents. 
Les nucleolites, une fois produits, grossissent et, d’homog^nes 
qu’ils etaient d’abord, peuvent offrir & leur tour la differencia- 
tion d’une couche corticale et d’une zone centrale et faire office 
de producteurs nouveaux . . . j’ai de bonnes raisons de penser 
qu’a ce moment tous les nucleolites produits sont de taille 
sensiblement dgale et qu’ils paraissent tous homogenes. . . . 
Je n’ai pas vu ce que deviennent ces fragments du nucleole, 
quelque soin que j’aie mis h scruter leur destinees. Je suppose 
que l’enveloppe du noyau se rompt, que les nucleolites mis en 
libertd gagnent par des mouvements propres la zone super- 
ficielle de la masse granuleuse pour s’y diviser activement. . . . 
Si ma hypothese etait fondle, le corps nucleolaire mis en 
liberte dans le plasma du kyste representerait en realite les 
debris de la fortune d’un noyau; ce serait le noyau lui-meme, 
segmentd, morceie, et le nom employe, celui de nucleoles, serait 
compietement impropre.” 

Weismann (’83), ova of Hydromedusae: in all the genera 
studied there is always a single large nucleolus, which some¬ 
times contains one or several vacuoles. 


1884. 

Ayers (’84) germinal vesicle of Oecanthus niveus: in smaller 
eggs a single nucleolus, in larger ones several; these nucleoli 
he considers as “nodules of nuclear filaments.” 


296 


MONTGOMERY. 


[Vol. XV. 


Carnoy (’84) distinguishes three kinds of nucleoli: ( 1 ) “nucle- 
oles nucleiniens,” which are parts of the chromatin network; 
( 2 ) “ nucRoles-noyaux,” which contain all the elements of a 
normal nucleus (namely, a membrane, chromatic filament, and 
nucleolar substance), while the substance in the remainder of 
the nucleus is allied to cytoplasm ; such nucleoli occur in 
Gregarines, large Radiolaria and Rhizopoda , Spirogyra, the asci 
of lichens, testicle cells of Littobius , and eggs of Pleurobrachia , 
Ascidia , and Nephthys; ( 3 ) “ nucleoles plasmatiques,” which 
contain no chromatin, but consist of a plastin network in 
which an albuminous enchylema is imbedded. 

Frommann (’84) studied fresh ganglion cells from the anterior 
horn of the medulla of the ox; their nucleolus shows “ eine 
Zusammensetzung aus feinen und derberen Kornchen und aus 
sehr kurzen Faden, mitunter auch einen netzformigen Bau mit 
theils ganz engen, theils etwas weiteren Maschen.” In the 
ganglion cells (of the ganglion Gasseri) of the rat, the nucleolus 
is usually homogeneous, as are those of the sympathetic ganglion 
cells of Bufo. 

R. Hertwig (’84), Actinosphaerium; in the resting nucleus 
there is one central nucleolus which consists of deeply staining 
nuclein and faintly staining paranuclein. The nucleolus is 
rarely spherical; when so, it consists mainly of nuclein, except 
for a small portion of paranuclein superimposed on the margin. 
In other cases the larger nuclein portion is of a curved dumb¬ 
bell shape, and “ gleichzeitig bildet das Paranuclein ein schwach 
gekrummtes Stabchen, dessen Kriimmung zur Krummung der 
Nucleinmasse senkrecht gestellt ist.” The connecting portion 
of the dumb-bell may disappear, “ so dass sich zwei Nucleoli 
bilden, welche von einander durch ein queres Stabchen Para¬ 
nuclein getrennt werden. . . . Hiermit beginnen die plurinu- 
cleolaren Kerne, wie sie fur gewohnlich bei Actinosphaerium 
beobachtet werden.” In most nuclei there lies a mass of from 
six to twenty nucleoli, which are smaller as they become more 
numerous: “ Hier ist es sehr schwer festzustellen, was aus dem 
Paranuclein geworden ist, und . . . bin ich zu dem Resultat 
gekommen, dass es als ein Korn im Centrum des Haufens von 
Kernkorperchen ist, dass es mit einem Fortsatz an jedes 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


297 


derselben herantritt und alle somit unter einander zu einer 
Rosette vereinigt. . . . Die staubformigen Nucleoli sind 
ursprtinglich vorhanden, erst allmahlich vereinigen sie sich 
zu grosseren Stricken, bis endlich nur ein einziger Nucleolus 
und Paranucleolus gegeben ist; dann tritt die Theilung ein.” In 
the resting nucleus all the chromatin is contained in the larger 
nucleolus. 

Jijima (’ 84 ) found that there are one or several nucleoli in 
the ripe eggs of Triclad Turbellaria , but none in younger 
germinal vesicles. 

Korschelt (’ 84 ), following Balbiani (’8i) and Leydig (’ 83 ), 
investigated the interesting structures in the cells of the 
salivary gland of Chironomus. The form and number of the 
nucleoli is mainly such as was described by Balbiani, “ meist 
aber sind sie ausgehohlt und von der Form einer mit sehr 
dickem Boden versehenen Schale. . . . Die Convexitat der 
Schale richtet sich immer nach der zunachst gelegenen Aussen- 
flache des Kernes. . . . Der Kernkorper besteht aus einer 
feinkornigen Masse, in welcher Vacuolen auftreten. . . . Von 
den Vacuolen fliessen oft einander benachbarte zu einer gros¬ 
seren zusammen.” The cross-striated structures described by 
Balbiani are not to be seen in the fresh nucleus, but, as noted 
by Leydig, first appear after the nucleus has remained under 
the microscope for some time; thus they may be possibly prod- , 
ucts of coagulation. “ Dass sie sich, wie dies Balbiani zeichnet, 
mit ihren fransenartig gebildeten Enden an die sog.' Kern- 
membranen anheften oder dass (nach Leydig) Anheftungsfaden 
von ihrer Oberflache zur Umgrenzung des Kernes hingingen, 
habe ich allerdings nie bemerken konnen. . . . Ich muss nach 
meinen Befunden . . . sagen, dass die “ Querstreifung ” der 
Bander auf einer Faltung ihrer Oberflache beruht und dass eine 
Zusammensetzung aus verschiedenartigen Schichten nicht vor¬ 
handen ist.” Further, Korschelt did not observe the envelop¬ 
ing membrane of these structures, described by Balbiani, though 
he corroborates the observation of this author that the end of 
the band gradually fuses into the mass of the nucleolus. From 
experiments on starving larvae, he concludes: “Es'scheint 
demnach das eigentliche Chromatin nicht die ganze Masse der 


298 


MONTGOMERY. 


[Vol. XV. 


Bander auszumachen, sondern nur einen Bestandtheil derselben 
zu bilden, der bei mangelhafter Ernahrung der Gewebe zuerst 
schwindet.” 

Lang (’ 84 ) remarks of the egg cells of Poly clad Turbellaria: 
“Das Kernkorperchen oder der Keimfleck ist stets als ein 
kugliger, relativ sehr grosser, intensiv gefarbter Korper zu 
unterscheiden.” 

Vejdovsky (’84) noticed a single nucleolus in the eggs of 
Oligochaeta. 

Wielowiejski (’84) studied the egg cells of various Arthropoda . 
In the Araneina and Acarina the larger nucleolus contains a 
single large or several smaller vacuoles, though no pulsating 
or amoeboid movements were noticed (in opposition to the 
observations of Balbiani). In Drassus and Lycosa there is a 
small mass of granules in place of a germinal spot; in Oniscus , 
a single large nucleolus; in Astacus, numerous peripheral ones; 
and in Musca, a large, irregularly spherical one. (He notes 
that the germinal vesicle differs from all other nuclei in that 
its contents do not stain at all, or only faintly, with acetic acid 
methylen-green solution.) 

Will (’84) studied in life the eggs of Bufo and Rana. Larger 
and smaller nucleoli may be distinguished; the latter increase 
somewhat in size, but never attain the dimensions of the 
preceding. Those nucleoli, then, which lie close to the nuclear 
membrane cause small protuberances (“ Knospen ”) of this 
membrane, each such bud next breaks off from the nucleus, and, 
still enclosing a nucleolus within itself, wanders towards the 
periphery of the cell, and there becomes a “ Dotterkern,” the 
disintegration of which furnishes the yolk granules. 

1885. 

Van Bambeke (’85) reviews the opinions of the following 
writers in regard to the nature of nucleoli: Flemming (’82), 
Strasburger, Pfitzner (’81), Retzius (’81), Leydig (’83), Balbiani 
(’81), Korschelt (’84), R. Hertwig (’84), Van Beneden (’83), 
Frommann (’84), Carnoy (’84), Brass, Wielowiejski (’84), and 
Rabl (’84). Nucleoli are rarely absent, and hence they must 
be regarded as an essential element of the nucleus. “Le 


LtfC. 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


299 


mode d’origine des nucleoles gendralement admis explique le 
rapport de ces elements avec la charpente nucleaire. . . . 
Flemming est dans le vrai en disant que si les nucleoles sont 
generalement suspendus au reticulum , ils ne sont pas en con¬ 
tinuity de substance avec ce dernier, mais constituent des 
elements speciaux. . . . Nous croyons devoir rapprocher du 
nucleole principal la formation recemment designee par Ed. 
Van Beneden, sous le nom de corpuscule germinatif, et plusieurs 
de celles appelees par Carnoy nucUoles-noyaux The nucleoli 
are probably reservoirs for masses of chromatin. 

Van Beneden and Julin (’85) found, in contradiction to 
Roule, that in the ovum there is only a single large “corpuscule 
germinatif ” in Clavelina , and neither smaller nucleoli nor any 
migration of nucleoli into the cytoplasm. 

Biitschli (’85), Ceratium tripos: most of the nucleoli of the 
individuals examined contained no nucleoli; only occasionally 
are one or two present, and then these evince a honey-combed 
(“wabige”) structure. In many Flagellata there is no trace of 
a nucleolus. 

Carnoy (’85) amplifies his observations of the preceding 
year, in which he had distinguished the following four types 
of nucleoli: ( 1 ) “nucleoles nucleiniens”; ( 2 ) “nucleoles plas- 
matiques”; ( 3 ) “nucleoles mixtes” (“qui sont constitues 
par la reunion des deux especes pr^c^dentes en un corps 
unique”); ( 4 ) “ nucleoles-noyaux.” Types 1 , 3 , and 4 are 
closely related, and all are sharply demarcated from type 2 . 
The “ nucldoles plasmatiques ” are plasmatic, albuminoid accu¬ 
mulations, and not chromatin material in reserve (in opposition 
to the views of Heuser, Guignard, and Pfitzner) : “Nous pre- 
fdrons dire qu’ils concourent avec les autres elements plas¬ 
matiques du noyau a l’elaboration du fuseau, dont les filaments 
constituants sont formds d’une substance, ou de diverses 
substances, presentant beaucoup d’analogie avec la plastine.” 
The “nucleole nucleinien ” may be composed of amorphous 
masses or of a skein of chromatin (the latter is the case in 
the testicle cells of Chilopoda , ova of Pleurobrachia and 
Cymbulia ) : “ Le nucleole central de beaucoup de cellules 
ganglionnaires est de nature nucleinienne et presente souvent 


300 


MONTGOMERY. 


[Vol. XV. 


la meme constitution filoi'de”; and similar nucleoli occur in the 
Protista and in various cells of the Arthropoda . The “ nucleole- 
noyau ” of the eggs of Cymbulia and Lithobius has a fine 
external membrane and a convoluted chromatin filament. In 
the amitotic division of the capsular ovarial cells of Gryllotalpa 
the nucleolus (formed of a central portion of chromatin and a 
peripheral layer of plastin) divides first so that each daughter- 
nucleus receives one nucleolus. But in the amitosis of the 
intestinal cells of Aphrophora the “nucleoles plasmatiques ” do 
not divide; and in the testicle cells (“metrocytes ”) of Scolo- 
pendra there is also a “nucleole plasmatique,” and at the 
commencement of the mitosis the “ nucleole se liquefie pour 
enrichir le caryoplasma,” and is not to be found later. The 
amitotic division of the fat cells of Geotrupes is introduced by 
a division of the “ nucleole-noyau.” 

Frenzel (’85) studied the cells of the mid-gut in insects at 
various stages of development. Bombyx dispar , larva: one 
large nucleolus containing a vacuole, in which lies a small 
spherical “ Nucleollolus.” Tachina, larva: here is one large 
nucleolus, “mit kurzen zackigen Auslaufern. In seinem 
Innern umschliesst er fast stets wenigstens einen, in der Regel 
aber mehrere, etwa 6 bis 12 , kugelige oder matt aussehende 
Gebilde, welche nicht gerade den Eindruck von festeren 
Korpern, sondern vielmehr von Vakuolen machen.” In cylin¬ 
der and gland cells of various insect larvae the nucleus is 
filled with a homogeneous fluid, “ in welche sowohl echte 
Nukleolen, wie auch nucleolenartige Korper (‘ Keimflecken * 
oder * Nukleolide ’) einerseits und andererseits zahlreiche ver- 
schieden angeordnete sehr klein aber stets gleich grosse 
Kornchen eingelagert sind, die hier * Kerngranula * oder 
‘ -granulationen ’ heissen mogen.” 

Leydig (’85) noticed in ganglion cells of Astacus a large, 
spherical, granular nucleolus, in which is a large cavity ; this 
nucleolar cavity stands in communication with that of the 
nucleus itself. We read further : “ Die Korper im Kern, die 
man Nukleoli nennt, sind Bildungen verschiedener Art” ; some 
arise out of the nodal points of the nuclear network, others out 
of the “ Kernplasma.” 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 301 

Rabl (’85) studied mitoses in cells of the larva of Salatnandra , 
and found that in the prophases of mitosis the nucleoli gradu¬ 
ally vanish and take part in the production of the chromatin 
threads. In the unripe germinal vesicle of Proteus , on the 
inner surface of its membrane, “ sieht man in unregelmassigen 
Abstanden von einander kugelige, stark glanzende, wie Oel- 
tropfen aussehende Korperchen,” which he assumes are neither 
nucleoli nor masses of true chromatin. 

Will (’85) studied the ovogenesis of Notonecta and Nepa. 
The young “Ooblast” contains one nucleolus bounded by a 
membrane and surrounded by smaller “ Chromatinballen” ; 
subsequently the latter bodies fuse together and form a closed 
ring around the nucleolus. The nuclear division of the ooblast 
is an amitotic one, and is preceded by a division of its nucleolus ; 
in each daughter-nucleus, then, the divided half of the primitive 
nucleolus breaks up into fragments, which become distributed 
through the nuclear sap, and the daughter-nucleus produces a 
new nucleolus without the aid of these particles. When the 
ovum proper is ripe, the nucleolus finally disappears. 

1886. 

Van Bambeke (’86) found that in the germinal vesicles of 
Aracfmida , Isopoda , Hymenoptera , and Meconema , the nucleoli 
and the chromatin do not stain with methylen green (corrobo¬ 
rating Wielowiejski) though they stain with carmine and 
haematoxylin ; “ Rien ne s’oppose, me semble-t-il, a ce que Ton 
considere le corpuscule germinatif comme etant equivalent a 
l’ensemble de la charpente chromatique des noyaux ordinaires 
[somatiques].” He concludes that there is no proof of the 
identity of the true nucleoli of the somatic cells with the ger¬ 
minal spots of egg cells. Two stages in the formation of the 
nucleolus may be distinguished in the ova of various Arachnids 
(Lycosa, Amaurobius , Argyronecta , Tegenaria , Attus, Theridium , 
Epeira , Zilla, Phalangium ): ( 1 ) there is a single large nucleolus 
(sometimes accompanied by smaller accessory ones), in which 
at first a few vacuoles arise, which later fuse to produce a single 
voluminous vacuole; and ( 2 ) the nucleolus becomes replaced 
by a mass of fine granules. In the ovarial egg of Amaurobius 


302 


MONTGOMERY. 


[Vol. XV. 


ferox the nucleolus consists of (i) a peripheral, less deeply 
staining portion ; and ( 2 ) of a more deeply staining and more 
highly refractive central portion, in which one large and several 
smaller vacuoles lie: “ Chose remarquable dans la vacuole 
centrale se voyait, a l’dtat frais, un granule fonce, doud d’un 
mouvement tres vif”; in this germinal vesicle a small, finely 
granular nucleolus is also present. Amoeboid movements of 
the germinal spot of Periplaneta were noticed. In the egg 
of Zilla there are from one to three homogeneous, spherical 
nucleoli, as also a large “tache principale ”; the latter is com¬ 
posed of two or three different substances, somewhat as in 
Amaurobius. 

Carnoy (’86), egg of Spiroptera strumosa: there is one large, 
central “nucleole nucleinien,” sometimes also one or two small 
“nucleoles plasmatiques ”; the former nucleolus is the only 
part of the nucleus which stains deeply with methyl green; 
it is bounded by a fine membrane, and contains eight “ baton- 
nets ” (chromosomes), so that it is comparable to a “nucleole- 
noyau.” Nematode from the stomach of Scyllium canicula: in 
the “oeufs tr&s jeunes . . . le filament nucleinien y est assez 
puissant, il parait continu. . . . Nous n’avons pu voir s’il se 
scindait d’abord en trongons ; nous croyons plutdt qu’il se 
localise par le retrait de ses anses, pour constituer un nucleole 
nucleinien pelotonne. Ainsi nait la tache de Wagner. Elle 
est toujours simple; elle se colore peu par le vert de methyle ”; 
no “ nucl6oles plasmatiques ” are present in this nucleus. In 
the egg of Filaroides mustelarum one or two “ nucleoles plas¬ 
matiques” occur; but in that of Ascaris lumbricoides such 
nucleoli are usually absent, and the chromatic filament extends 
through the whole nucleus. In Ascaris sp. (from the dog) there 
is one “nucleole plasmatique” in young eggs. 

Heathcote (’86) noticed in the egg of Julus one nucleolus with 
vacuoles ; it disappears before the production of the pole bodies. 

Knappe (’86), ovarian ova of Bufo: The nucleoli show amoe¬ 
boid movements in life, and these movements probably lead to 
the dissolution of the nucleoli, by causing the latter to first 
break into fragments, these fragments afterwards dissolving in 
the nuclear sap. 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


303 


Pfitzner (’86a) distinguishes in the nucleus: “ Das Achroma- 
tin, eine geformte farbbare Substanz, das Chromatin (mit der 
Unterart der Nucleolensubstanz, des Procbromatins) und eine 
geformte nicht farbbare Substanz, das Parachromatin.” In a 
second paper (’86b) he studied Opalina: here are several 
nucleoli flattened against the nuclear membrane; “bei der 
Kinese verschwinden sie allmahlich, aber spater als bei anderen 
Objekten bisweilen sind sie noch bis zur Metakinese vorhan- 
den.” Though they are occasionally found at the poles of the 
spindle they take no part in the formation of the chromatin 
elements, and in the daughter-nuclei reappear at a distance 
from the latter elements. For denoting the substance of the 
nucleoli he substitutes for his earlier term “ Prochromatin ” 
the term “ Pseudochromatin,” since “ das Chromatin und die 
Nucleolensubstanz wohl nichts Anderes mit einander gemein- 
sam haben, als die untergeordnete Eigenschaft, sich bei den 
meisten Farbemethoden gleicherweise stark zu farben.” 

Platner (’86) investigated the ovogenesis of Avion and Helix. 
In Avion there appears first in the ‘‘primitives Ei” a small, 
completely spherical nucleolus, to which he limits the name 
“Nucleolus”; “weiterhin enthalt das Keimblaschen den 
eigentlichen Keimfleck. Dieses ist zu Beginn seines Auftre- 
tens meist rundlich mit hervorspringenden Erhabenheiten, als 
sei er durch Contraktion eines Knauels entstanden. Zuweilen 
erscheint er auch mehr ringformig oder ganz unregelmassig. 
Immer aber verdichtet er sich bald zu einem vollig runden 
homogenen Element, welches Kernfarbstoffe begierig auf- 
nimmt und den Nucleolus bedeutend an Ausdehnung iiber- 
trifft.” (His figures show the two to be in close contact.) A 
number of clear vacuoles begin to appear in the “ Keimfleck ”: 
“ Sie sind rund und von verschiedener Grosse . . . und schei- 
nen nur dazu zu dienen, weitere Veranderungen einzuleiten. 
Sie verschwinden namlich alsbald wieder, und in dem stetig an 
Grosse zunehmenden Keimfleck scheidet sich mit wachsender 
Deutlichkeit eine heller gefarbte und eine dunklere Partie. 
Letztere, dem “corpuscle germinative” van Benedens ent- 
sprechend, ist von geringer Ausdehnung, rundlich oder lang- 
lich oval und liegt excentrisch in der von runden Contouren 


304 


MONTGOMERY. 


[Vol. XV. 


begrenzten hellen Substanz, die demnach auf dem Querschnitte 
halbmondformig erscheint. Sie diirfte dem von van Beneden 
als “ prothyalosome ” bezeiehneten Gebilde entsprechen. Es 
sei mir daher gestattet, sie Hyalosoma zu benennen. In vollig 
entwickelten Eiern ist dieses Element nahezu vollig farblos 
und erscheint aus feinen Kornchen zusammengesetzt. Die 
gefarbte Partie des Keimflecks tritt dadurch um so scharfer 
hervor, man kann sie im Anschluss an van Beneden Keim- 
kdrperchen nennen.” The nucleolus of the ripe egg “ liegt 
excentrisch und besteht wieder aus dem runden zart granulirten 
Hyalosoma, sowie in dem peripher in demselben gelagerten 
Keimkorperchen, welches sich stark farbt und keine weitere 
Differenzierung erkennen lasst. Dem hellen Hyalosoma meist 
dicht anliegend findet sich der intensiv sich farbende Nucleolus 
oder der kleinere Keimfleck.” Platner considers that by the 
last division of the ovocyte the “ Nebenkern ” disappears and 
becomes a constituent of the nucleus. “ Bei Ausbildung der 
Furchungsspindel konnte ich mit Sicherheit constatiren, dass 
die Spindelfasern aus der unfarbbaren Substanz des Eikerns 
hervorgingen. Diese ist bei sich entwickelnden Eiern im 
Keimfleck enthalten, in welchem sie sich bald als Hyalosoma 
differenzirt.” In Helix the “ primitive Eier . . . entbehren 
des schonen grossen Nucleolus. . . . Daher enthalt ihre defi¬ 
nitive Form auch nur einen Keimfleck, welcher weiterhin 
dieselben Veranderungen zeigte wie bei Avion!' It may be 
noted in conclusion that in the spermatogonium of Avion the 
nucleolus appears in the nucleus at the same time that the 
“ Nebenkern ” appears in the cytoplasm. 

Schauinsland (’86) found one or two large nucleoli in the egg 
of Bothviocephalus vugosus. 

Stuhlmann (’86) investigated the early stages of the ovum in 
a large number of species, more particularly of the Avthvopoda. 
Cavabus memovalis : there are numerous “ chromatin ” granules 
in the young eggs, which increase in number and size; later a 
granular nucleolus appears : “ Es ist schwer zu entscheiden, ob 
der Haufe von chromatischen Kornern zu einem grossen Ballen 
zusammenschmilzt, oder ob sich einer, wohl das urspriinglich 
central gelegene, zum Nucleolus ausbildet oder endlich ob 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


letzterer eine ganz neue Bildung ist. . . . Wenn aber schon 
Dotter ausgeschieden ist, hat der Nucleolus fast stets eine 
Form, die aufs Tauschendste einer Eichel gleicht. . . . Wir 
sehen an dem Nucleolus einen helleren, vollig homogenen 
Theil und einen dunkler gefarbten, welcher fein granulirt ist 
und wie mit einer Menge von winzigen Vacuolen durchsetzt 
erscheint. Dieser dunklere Theil umgreift wie die Cupola 
einer Eichel den helleren Theil. Um die Formahnlichkeit 
ganz zu vollenden, sitzen haufig auf der Kuppe der homogenen 
Halfte noch einige dunkle Kornchen. . . . Auf einem Aequa- 
torialschnitt sieht man nun, dass der dunklere Theil eine Zone 
um den helleren Theil bildet.” This enormous nucleolus 
measures 67 /x; it disappears when the nucleus wanders to 
the periphery of the egg. Carabus auratus and P*terostichus 
elatus: one spherical nucleolus, containing a few small vacu¬ 
oles, and its size increases with that of the nucleus; later it 
assumes a peripheral position, “und in seiner Nahe treten mehr 
oder weniger kleine Chromatinkugeln auf, wahrend der Nucle¬ 
olus selbst kleiner zu werden scheint ”; the nucleolus dis¬ 
appears, then the small “Kugeln” unite to form a larger 
spherule, and finally the latter also vanishes. In the egg of 
Dytiscus marginalis there are no true nucleoli, only irregular 
masses of chromatin. Egg of Silpha : one granular nucleolus, 
which increases in size up to a certain point, and later, when 
vacuoles arise in it, a number of small spherules become appar¬ 
ent outside of the nucleolus: “ Ob dieselbenaus dem Nucleolus 
stammen oder ob sie als Paranucleolen des Kerngeriistes auf- 
zufassen sind, weiss ich nicht,” though he does not think that 
they are products of the nucleolus; the nucleolus, as well as a 
portion of the nucleus, disappears later. Necrophorus vespillo : 
several non-homogeneous germinal spots, later a single nucleolus, 
which finally vanishes. Eggs of Geotrupes and Cetonia : several 
small, spherical or elongated nucleoli, which occupy a central 
position in the nucleus, and increase in number and size; 
“ Dieselben liegen in concentrischer Anordnung um einen 
homogenen Kern, der etwas dunkler als die Kerngrundsubstanz 
gefarbt ist.” Lina popnli: at first there is one large and one 
small nucleolus; in this stage “ sind im ganzen Keimblaschen 


3°6 


MONTGOMERY. 


[Vol. XV. 


mit Ausnahme der Randzone ganz feine klare Blaschen ver- 
theilt, welche ich jedoch als Kunstprodukte ansehen mochte ”; 
later there lies in one part of the nucleus a group of minute 
nucleoli; then a portion of the nucleus breaks off and wanders 
into the cytoplasm, while the remaining portion of the nucleus 
retains one small nucleolus; and lastly, when the nucleus 
becomes amoeboid in shape, it contains one large vacuolated 
nucleolus, “sowie mehrere kleinere chromatische Korper.” 
Lycus aurora: at first there is no nucleolus, later a large and 
a small one (both granular); when the nucleus wanders to the 
periphery of the egg it retains one of the nucleoli, which 
subsequently disappears at the same time as the nucleus 
does. Periplaneta orientalis: at first there is no nucleolus, 
“ derselbe bildet sich erst allmahlich heraus. . . . Wir sehen 
ausser dem etwas kornigen Nucleolus eine Anzahl kleinerer 
stark farbbarer Kugelchen, die wohl als Bestandtheile des 
Kerngeriistes, als Paranucleolen aufzufassen sind.” Gryllotalpa 
vulgaris: in the immature egg “ ein eigentlicher Keimfleck ist 
nicht vorhanden; vielmehr liegen in der Kerngrundsubstanz 
zerstreut Chromatinpartikel von 4/1 Durchmesser bis zu 
unmessbarer Feinheit”; when the nucleus has assumed a 
peripheral position a large nucleolus is produced in it, “ wohl 
durch Verschmelzung mehrerer kleinerer.” Locusta viridissima: 
in maturer ova a large but lightly staining nucleolus, “ von dem 
aus ein Kernnetz seinen Ursprung nimmt.” Pieris brassicae : 
one large, homogeneous germinal spot, which later acquires 
vacuoles and divides into three parts. Sphinx ligustris: in the 
immature germinal vesicle lies a large, excentric nucleolus, con¬ 
taining vacuoles; “ ausser letzterem finden sich noch einige 
wenige Paranucleolen ”; at the time when the nuclear frag¬ 
ments break off, the nucleolus becomes paler and then vanishes. 
Zygaena filipendulae: at first no nucleolus is present, later 
there is a larger one with vacuoles, as well as a smaller one, 
“ der sich wohl von dem grossen abgelost zu haben scheint ” ; 
subsequently both disappear. Musca vomitoria: there is at 
first in the germinal vesicle a single, large, excentric nucleolus, 
but later appear in it “ eine Anzahl von Paranucleolen und ein 
Nucleolus, . . . von welchen letzterer aus einem Haufchen von 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES . 307 

kleinen, gefarbten Kiigelchen besteht.” In the egg of Anabolia 
there is one large nucleolus, but in those of Vespa germanica 
and V. media apparently no true nucleoli are present. There 
is a large granular nucleolus in the larger germinal vesicles 
of Bombas terristris. Trogas lutorius: there is one large, 
irregularly shaped nucleolus and two smaller ones; all these 
finally disappear, and their place is taken by smaller granules. 
Banchus falvipes: at first no nucleolus is present, later one or 
three large nucleoli appear, but all of them vanish subsequently. 
In the egg nucleus of Pimpla sp. only a number of small 
granules are to be found, and at a later period still smaller 
granules. Anomalon circumflexum: in the youngest germinal 
vesicles no nucleolus is to be found, in older ones there is 
a single large one; this has nothing to do in the formation of 
the “ Dotterkerne,” and disappears when the nucleus does. 
There is one spherical germinal spot in Ophion ventricosum } 
but not in O. lateum. Ephialtes liturater: in the smaller nuclei 
a considerable number of “ chromatic ” bodies occur, while in 
the older ones there is a single large nucleolus. Ambyteles 
castigator: one large nucleolus, in older ova also several smaller 
ones. Epeira diademata: here is one large spherical nucleolus, 
which later becomes jagged in outline and evinces vacuoles, 
which may unite to produce a single larger vacuole : “ In sel- 
tenen Fallen kann man einen Zerfall des Nucleolus in mehrere 
kleinere sehen, was jedoch wohl eine pathologische Erscheinung 
sein diirfte.” Glomeris marginata: one large, spherical or 
angular nucleolus, and later also a smaller one : “ Hochst 
wahrscheinlich stammt dieser von dem grossen Nucleolus ab ” ; 
the smaller nucleolus disappears subsequently. In the egg of 
Peripatus edwardsii one nucleolus forms itself gradually, and 
vacuoles begin to appear in it. In Amaroecium rubicundum a 
single large nucleolus is present; while in Clavelina lepadi- 
formis the nucleolus is probably formed out of the central 
chromatin masses. From these numerous observations Stuhl- 
mann draws the conclusion: “ Aus Allem schien mir hervorzu- 
gehen, dass das Schwinden des Nucleolus nicht zum Wesen 
der Eireifung gehort, besonders weil ich ihn bisweilen (so bei 
Silpha) so lange verfolgen konnte, als noch ein Rest des 
Keimblaschens im Ei sichtbar war.” 


30S 


MONTGOMERY. 


[VOL. XV. 


Vigelius (’86) finds in the egg of Bngula one large nucleolus, 
containing vacuoles. 

Will (’86) studied the maturation of the egg of Colymbetes. 
“ Dem Kernkorperchen oder Nucleolus . . . kann nach meinen 
Untersuchungen keinerlei morphologische Bedeutung zukom- 
men. Was wir Kernkorperchen nennen, ist nach meiner 
Auffassung nichts als ein besonders grosses Stuck Chromatin- 
substanz. So konnen wir es verstehen, dass bald eines, bald 
mehreres, bald gar keine vorhanden sind.” 

1887. 

Boveri (’87): in the ovum of Ascaris megalocephala bivalens 
there are no true nucleoli when the tetrads are formed. In the 
variety univalens there is usually one “ achromatisches kugeliges 
Korperchen. Von dem “ Prothyalosoma,” das an den van 
Beneden’schen Eiern den Keimfleck [Vierergruppe] umgiebt 
und welches im weiteren Verlauf bei ihm eine so grosse Rolle 
spielt, habe ich weder auf diesem Stadium, noch spater die 
geringste Spur wahrgenommen.” 

Eisig (’87) remarks in regard to the egg of Capitellids: “Der 
urspriinglich rundliche, jederzeit durch Dichtigkeit und hohes 
Tinctionsvermogen auffallende Keimfleck erleidet im Laufe 
seines Wachsthums offenbar Theilungen; denn man findet ihn 
in spateren Stadien mit ein oder zwei verschiedengradig abge- 
schnurten Kuppen besetzt; ausserdem trifft man schon friihe 
mehrere Pseudonucleoli, welche offenbar Produkte des Haupt- 
nucleolus darstellen, in dem Keimblaschen zerstreut.” He 
notes, further, that in the maturing ovum the nucleolus does 
not increase in size in equal proportion to the size of the 
nucleus. (To judge from his figures, the nucleoli are not 
homogeneous.) 

Fraipont (’87) found in the germinal vesicle of Polygordius 
several nucleoli of unequal size. 

Henking (’87) studied the eggs of Phalangids. In the ovarial 
egg a sickle-shaped body lies at one pole of the nucleolus: “es 
scheint, als wenn in ihm und dem Keimfleck die Chromatin- 
substanz des Keimblaschens sich koncentrirt hatte.” In the 
nearly ripe egg there is one large nucleolus, which is not homo- 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


309 


geneous, and a number of smaller globules, these latter stain¬ 
ing as the former, and some of them containing vacuoles: “ sie 
stellen einerseits eine Zusammenballung der bisher ganz unre- 
gelmassigen, im Keimblaschen vertheilten Chromatinsubstanz 
dar, ruhren andererseits aber wohl vom Keimfleck her.” These 
bodies have all disappeared in the ripe egg. 

Hubrecht (’87) noticed only a single nucleolus in the egg of 
Cerebratulus sp.; as to the egg of Pelagonemertes , he figures 
one nucleus containing one large and several smaller nucleoli, 
and another nucleus with only numerous small nucleoli. 

Kosinski (’87, ’93, mentioned by Lavdowsky, ’ 94 ): within the 
nucleolus of cancerous cells there is sometimes a vacuole, and 
within the latter a small body which Kosinski considers may 
correspond to Carnoy’s “ nucleoles-noyaux such nucleoli have 
the faculty of division, and of wandering through the nuclear 
membrane into the cytoplasm. 

Lukjanow (’87a), stomach epithelium of Amphibians: in the 
cytoplasm of the cylinder epithelium are structures of various 
form (“ Nebenkerne ”), which stain in general like the nucleoli. 
In some of the nuclei of the deep layer of gland cells each 
nucleolus is joined with a karyosome. 

Lukjanow (’87b) distinguishes three kinds of nucleoli in 
muscle cells of Vertebrates: ( 1 ) “ Plasmosomen ”; ( 2 ) “ Karyo- 
somen”; (3) “ Kernkorperchen von gemischtem Charakter.” 
The first stains deeply red (eosin), the second blue-violet (haema- 
toxylin), while the third stains a mixed color with these two 
stains (when used together). He remarks also: “ dass in 
manchen Kernen die Kernkorperchen ganzlich fehlen, in 
anderen entweder nur eine Kategorie derselben, oder mehrere 
zugleich vertreten sind. . . . Zuweilen liegt das Kernkorper¬ 
chen sogar ganz ausserhalb des Kernes.” 

Nussbaum (’87) found in smaller eggs of Hydra a single large 
nucleolus, while in larger ova several are present. “ In frischem 
Zustande sieht man in den allerersten Stadien neben den Keim- 
flecken noch eine blasse Kugel, die im Gegensatz zu den Nucle- 
olen des Keimblaschens keine Farbstoffe in sich aufnimmt.” 

O. Schultze (’87) studied the maturation of the egg in Rana 
and Triton. In the unripe germinal vesicle there are larger 


3io 


MONTGOMERY. 


[Vol. XV. 


nucleoli near the nuclear membrane, and smaller ones at the 
center of the nucleus: “Dass sie sich durch Theilung vermehren, 
kann keinem Zweifel unterliegen, denn nicht nur sind dieselben 
in ganz j ungen Eiern grosser und weniger zahlreich, . . . 
sondern die grosseren Keimkorperchen weisen durch Ein- 
schniirung und Zerkliiftung auf eine Vermehrung durch Thei¬ 
lung hin.” He does not consider that such daughter-nucleoli 
are again capable of division, but that the process is rather a 
“ Losungsphanomen.” All the nucleoli are homogeneous, but 
vacuoles are produced in them by ^ per cent normal salt solu¬ 
tion. In larger ova a considerable number of nucleoli lie periph¬ 
erally, and there is also a central group of them; and, still later, 
the peripheral nucleoli commence to stain less intensely, and 
the greater number are centrally situated. The nucleus of the 
maturing egg consists of “ Membran, Kernsaft und Keimkor¬ 
perchen,” a chromatin network being absent; and the microsomes 
of the chromosomes are formed from the smallest, most centrally 
placed nucleoli. 


1888. 

Bohm (’ 88 ) found in the egg cell of a 5 cm. long Ammocoetes 
of Petromyzon a homogeneous nucleolus, “ an dem sich sehr oft 
eine kleine Vacuole zeigt, welche mit einer feinen Strasse bis 
an die Oberflache des Fleckes [nucleolus] reicht.” At the 
animal pole of the nucleus lies a disc-shaped mass (“Deckel”): 
“ rathselhaft ist die Bedeutung des Deckels.” (Compare the 
extranuclear structure found by Lukjanow, ’88.) 

Boveri (’88): in the female pronucleus is neither a prothyalo- 
soma nor a hyalosoma, such as were described by Van Beneden 
(’83); the hyalosoma is probably “ ein durch Schrumpfung 
entstandenes Artefakt.” Just before copulation “ zeigen sich 
die ersten Spuren achromatischer Kernkorperchen als ganz 
kleine Kornchen, die . . . stets ... in nachster Nachbarschaft 
der chromatischen Elemente sich finden, ... so dass die Ver- 
mutung nahe gelegt wird, dass sie sich aus diesen absondern.” 

Fiedler (’ 88 ) studied the egg development of Spongilla: one 
large homogeneous nucleolus is present in the germinal vesicle. 
In the nuclear division (which is intermediate between the 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


311 

mitotic and the amitotic) “ der gesammte sonstige — iibrigens 
sparliche — Chromatin inhalt des Kernes vereinigt sich . . . mit 
dem Kernkorperchen zu einem kugeligen Gebilde, und erst 
dieses zerfallt dann durch allmahliche Zerschniirung in zwei 
kleinere, unter sich gleich grosse Kernkorperchen, welche an 
die beiden Pole des Kernblaschens riicken.” 

Graff (’88) found in the egg of Spinther either a mass of 
granules or a single nucleolus; the nucleolus may be either 
granular or contain a large vacuole. 

R. Hertwig (’88): in nuclei occur “ chromatische ” nucleoli, 
and “das unter gewohnlichen Verhaltnissen nicht farbende 
Paranuclein, welches zumeist rundliche Korper, die Paranucleoli, 
bildet. Die Paranucleoli konnen entweder die einzigen Kern¬ 
korperchen im Kern sein (gewohnliche Gewebszellen, reifes Ei 
und Furchungszellen) oder sie finden sich neben den chromati- 
schen Nucleoli, unter Umstanden auch als Einschliisse derselben 
(Keimblaschen der unreifen Eier, Kerne von Actinosphaerien 
und anderen Protozoen) vor. . . . Zweifelhaft wird es dagegen 
gelassen, ob auch der Substanz des achromatischen Geriistes 
. . . nicht . . . vielleicht auch Paranuclein [ist], welches sich 
durch seine Anordnung von den Paranucleoli unterscheidet.” 
The centrosomes are probably derived from the paranucleoli, 
and the paranuclein is “die befruchtende Substanz” (these 
views have subsequently (’96) been retracted). 

Kultschitzky (’88) found in the youngest eggs of Ascaris 
marginata one “ Kernkorperchen,” which afterwards “ in zwei 
Stiickchen zerfallt, deren eines sich intensiv mit Karmin farbt 
und alle Eigenschaften des Chromatins bewahrt, das andere 
sich in die blasser gefarbte gewohnliche Kernkorperchen ver- 
wandelt ” ; the latter he terms the true “ Kernkorperchen,” 
which from this stage on gradually decreases in size, and finally 
disappears. 

Leydig (’88) gives the results of numerous comparative inves¬ 
tigations on germinal vesicles ; most of these observations 
were made cfn the living egg, fixing reagents having been little 
employed. Nephelis vulgaris: here there is one nucleolus, 
which sometimes has a long process, “in dessen Nahe kleine 
rundliche Ballen von gleicher Art, wie er selber ist, liegen, so 


3 12 


MONTGOMERY. 


[Vol. XV- 


dass man die Entstehung der letzteren durch Abschniirung 
von dem Fortsatz sich denken darf.” Argulus foliaceus: in 
young eggs there is one large nucleolus with clear spaces in it, 
showing that the nucleolus “aus Theilen besteht, die allmah- 
lich von einander weichen, so dass man alsdann in anderen 
Thieren anstatt eines Keimflecks eine ganze Anzahl kleinerer 
vor sich hat”; these nucleoli are often jagged in contour ; by 
treatment with chromo-acetic acid “ bekommen die Keimflecke 
eine Querzeichnung, so dass sie wie aus Querstiicken zusam- 
mengesetzt erscheinen.” Tetragnatha: one large nucleolus 
with dark contours, and several smaller pale, granular ones, 
which gradually disappear during the maturation of the egg. 
Lycosa : “ Ein einziger, grosserer Keimfleck zeigt sich . . . und 
dieser bietet das Bild eines Knauels dar.” Theridium : the large 
“Hauptkeimfleck hat die Beschaffenheit eines stattlichen, 
aus scharf geranderten kleinen Korpern zusammengesetzten 
Ballens. Von ihm nun weg zieht sich ein Strang solcher Kor- 
perchen oder Theilstiicke iiber die Grenze des Keimblaschens 
hinaus in den Dotter hinein. In einzelnen Eiern, deren gros¬ 
ser Keimfleck das Bild gewundener und geknauelter Faden 
giebt, konnen die kleinen Theilstiicke zusammenhangend oder 
in bereits abgelosten Gruppen abermals in den Dotter sich 
erstrecken. Ja ich glaube an dem lebenden Ei verfolgt zu haben, 
wie Theile der geknauelten Faden sich zu einzelnen Ballen 
zusammenschoben und in den Dotter vordrangen ”; there are 
present also one or several pale “ Nebenkeimflecke.” Phalan- 
gium: the young ovum has one large nucleolus containing 
vacuoles ; “ Wiederholt habe ich beobachtet, dass ein solcher 
Keimfleck — das lebende Ei mit Mundspeichel befeuchtet — 
unter dem Mikroskop allmahlich verblasste und zuletzt fur das 
Auge vollig verschwand.” Lithobius: there may be one granu¬ 
lar nucleolus, or numerous nucleoli, each with a granular core : 
“Wieder eine andere Form ist die, dass die amobenartigen 
Gebilde in ihrem Innern einen hellen, kernartigen Fleck mit 
centralem Piinktchen zeigen und am Rande feinstrahlig sind”; 
in other germinal vesicles there may be present numerous 
small nucleoli, either irregularly grouped or arranged in “ kurze, 
goldrollenahnliche Saulchen . . .; ein andermal stosst man auf 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 313 

langere fadige Aufreihungen, deren Strange zu Schlingen gebo- 
gen oder geknickt sind.” In these ova two kinds of nucleoli 
may occur, namely, numbers of the small ones just described, 
and 3, large one with dark contours, which has a central 
vacuolar, granular portion, and is peripherally homoge¬ 
neous ; but nucleoli also occur which are intermediate between 
these two kinds. Geophilus electricus: here are numerous 
small, pale nucleoli and a large one, which has a finely granu -1 
lated core, and an outer homogeneous layer, the latter portion 
consisting of concentric layers; further, he noticed the infundib¬ 
ular structure first found by Balbiani on the outer surface of 
the nucleus, though he remarks that it is especially apparent in 
eggs in which post-mortem changes have commenced (!), and 
concludes : “ Wir haben es sonach beziiglich des Trichters mit 
einer Ausbuchtung jenes Hohlraumes oder Lichtung zu thun, 
welche von der Hohlung um das Keimblaschen herum in den 
Dotter dringt.” The basis of this infundibulum empties into 
a space around the nucleus, and not into the nucleus itself (as 
opposed to Balbiani’s observations); Leydig also thinks that 
particles of finely divided nucleoli penetrate separately out of 
pores which are present in the nuclear membrane, and that 
these particles, arrived in the cytoplasm, fuse together to form 
a large “Ballen.” Stenobothrus: in the ova of the proximal 
portion of the egg tube there are either numerous small nucleoli 
or a dense mass of very fine granules ; in riper germinal vesicles 
they are much larger and resemble somewhat the nucleoli 
in the salivary glands of Chironomus; masses of nucleolar 
substance wander out of the nucleus into the cytoplasm. In 
Pemphigus bursarius there is one compound nucleolus, with fine 
radiating processes ; and in Meloe violaceus there are numer¬ 
ous nucleoli, each of which has the structure of the single one 
of the preceding species. Gasterosteus aculeatus: in the month 
of May there are numerous germinal spots, sometimes densely 
grouped, sometimes arranged in rows ; the gradual thickening 
of the nuclear membrane takes place at the cost of nucleolar 
substance. Triton taeniatus: the germinal vesicle at the end 
of October contains numerous nucleoli of unequal size, many of 
which are arranged in columns ; the peripheral ones probably 


3 H 


MONTGOMERY. 


[Vol. XV. 


wander into the cytoplasm. Salamandra maculosa y larvae : 
the “ Urei ” has a single large nucleolus. Bnfo cinereus , larvae 
of several months : concludes “ dass die Keimflecke, wenn noch 
winzig klein, aus den Knotenpunkten des Spongioplasmas 
entstanden sind, und nachdem sie eine gewisse Grosse erreicht, 
die Form und Sonderung einer Amobe besitzen. Dieselben 
stellen sich jetzt dar wie hiillenlose, kleine Zellen, air. denen wir 
einen homogenen kornigen Korper, der feinzackig oder selbst 
in feine Strahlen ausgezogen ist, unterscheiden und im Innern 
einen lichten, kernahnlichen Fleck, in dem sich noch ein Kor- 
perchen abzeichnet”; numbers of such nucleoli may later fuse 
together, “ unter Vermittelung ihrer Zackenspitzen.” Rana 
esculenta: in the smallest ova there is only a single large 
nucleolus, with a vacuolar central portion and peripheral 
radiating strands ; in larger eggs there are a number of smaller 
nucleoli, each of which has the same structure as the primitive 
one ; Leydig believes that nucleoli wander out of the nucleus, 
since he found a granular mass on the outer surface of the 
latter. The ova of Sus scrofa , Myoxus nitela , and Talpa euro¬ 
pea contain each a single nucleolus. 

Lukjanow (’ 88 ) investigated the stomach mucosa of Salaman¬ 
dra. There are several, usually club-shaped nucleoli (“ Nucleoli 
claviformes”), the smaller, often funnel-shaped, end of which 
is in contact with the nuclear membrane. He concludes 
“dass die kolbenahnliche Form des Nucleolus . . . auf eine 
Vorbereitung zur Inhaltsentleerung hinweist. Der Kolben 
entleert seinen Inhalt etwa ebenso, wie die Becherzelle ihren 
Schleim entleert ” ; and he supports this conclusion with the 
observation that a mass is often found on the outer surface of 
the nuclear membrane which stains like the nucleolus. 

Nagel (’ 88 ) studied the human egg. The “ Primordial-Ei ” 
has a single nucleolus ; those which contain no nucleoli he 
believes do not develop further. In the ripe egg amoeboid 
motions were noticed in life (studied in liquor folliculi). 

Sanfelice (’ 88 ) terms the nucleolus of the spermatoblast 
“ nucleus,” and the nucleus, “cell.” What he calls the nucleus 
then divides karyokinetically (but that this process is a division 
of the nucleolus may be deduced from his figures 6 o and 62 ). 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 315 

Scharff (’ 88 ) studied the maturation of the eggs of various 
Teleosts. In the smallest ova examined (.011 mm.) there are 
numerous peripheral nucleoli, and a few which are central in 
position. In larger eggs (.03 mm.) “ the nucleoli show an 
inclination to gather still more towards the periphery of the 
nucleus . . . one or more of the nucleoli become larger than 
the others, and in their interior refractive specks are visible 
which have sometimes been described as endonucleoli.” In 
still larger ova (.08 mm.) “in some cases the big nucleoli dis¬ 
appear almost completely, leaving an unstained part around 
them.” In Conger he “noticed a small nucleolus being con¬ 
stricted off from a larger one.” He figures outside of the 
germinal vesicle of Gadus certain granules, and these he 
considers are emigrated nucleoli which are destined to become 
dissolved there, though he holds it possible that “some find 
their way to the surface of the egg to form the nuclei of the 
follicular epithelium” ; in eggs which have attained the 
dimensions of .132 mm. the nucleoli become very irregular 
in shape. In the Trigla egg of .13 mm. the surface of the 
nucleolus is raised into small protuberances, most of which 
contain a nucleolus ; these protuberances later break off and 
become the yolk spherules (in corroboration of Will, ’84). 

Schewiakoff (’ 88 ), Euglypha: the nucleolus gradually dis¬ 
appears in the prophasis of mitosis. 

Steinhaus (’ 88 ), intestinal cells of Salamandra; karyosomes 
and plasmosomes are distinguished within the nucleoli, and are 
usually combined in pairs with one another. Plurinucleolar 
nucleoli are formed by continued divisions of a single nucleolus, 
“et les nouveaux nucldoles s’dloignent Tun de l’autre, probable- 
ment a l’aide de mouvements amoeboides ou d’autres qui leur 
sont propres.” Plasmosomes when extruded into the cytoplasm 
increase greatly in size, though this increase is due to mere 
imbibition of some substance ; each such extruded nucleolus, 
combining with a karyosome, develops into a new nucleus. 

Vejdovsky (’ 88 ) studied the maturation of the egg of Rhyn- 
chelmis. The embryonal genital cells contain no true nucleoli. 
The nucleolus does not stain when it first appears (in very 
young stages). Subsequently it is always excentric in position, 


316 


MONTGOMERY. 


[Vol. XV. 


perfectly spherical, and consists of a central, homogeneous, 
deeply staining portion, and an outer unstaining envelope 
(judging from his Fig. 5, Tab. 3, I would consider this sup¬ 
posed envelope to be a vacuole in which the nucleolus lies). 
In the more advanced ovum this envelope has disappeared, and 
the nucleolus has increased in size, but is no longer homogene¬ 
ous, since it contains a number of deeply staining granules. 
When “das Kernkorperchen die oben angedeutete Grosse 
[.013 mm.] erlangt hat, beginnt es sich einzuschniiren, was 
gewiss auf dessen Theilung hinweist”; he believes that this 
division is rapid, “dass es aber thatsachlich so geschieht, 
beweist die Thatsache, dass in den reiferen Eiern in der Regel 
zwei Kernkorperchen vorhanden sind. Das neu entstandene 
Kernkorperchen liegt anfanglich in der Nahe des alteren und 
ist etwas kleiner als dieses; spater entfernt es sich mehr oder 
weniger und wachst zu der Grosse des ersteren heran.” In the 
ripe egg two nucleoli are present, or there may be three or four, 
the latter two having been divided off from the former; each 
of these consists of an inner chromatic portion and an achro¬ 
matic envelope ; the latter is porous, and “man kann voraus- 
setzen, dass durch die Poren die fliissige Nahrung in das Innere 
des Kernkorperchens eindringt.” When this envelope has 
vanished, each nucleolus is formed of (1) a hyaline, homogene¬ 
ous fluid, in which (2) a delicate network arises, the nodal 
points of which are represented by the previous granules of the 
nucleolus ; “ kurz und gut, die Kernkorperchen unserer Eier 
sind chromatische Kernfaden. . . . Die intensive Farbung 
sowohl der Knotchen als des Fadenwerkes erleichtert die Ver- 
folgung des metamorphosirten Kernkorperchens, welches jetzt 
ganz und gar den Kernen des spateren Blastomeren gleich- 
kommt.” (The descriptions do not enable one to determine 
whether all the nucleoli become thus metamorphosed.) 

Waldeyer in his “Referat” (’88) agrees with Klein “dass 
die Nucleolen nur stark verdickte Knotenpunkte des Netz- 
werkes der Geriistfaden [chromatin], also mit den letzteren 
identisch seien. . . . Die Bedeutung aller dieser Dinge fur das 
Zellenleben ist noch fast vollkommen dunkel.” 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 




1889. 

Bergh (’ 89 ), Urostyla: the fragments of the macronucleus 
contain true nucleoli, while the micronuclei do not. 

Brass (’ 89 ) states: “ Fur gewohnlich erscheint jedes Kernkor- 
perchen rund, sehr haufig kugelrund; es besteht entweder aus 
einer gleichartigen Masse oder es sind in derselben einige 
hellglanzende Kornchen ausgeschieden, oder aber es finden sich 
in ihm dichtere, weniger glanzende Kornchen. ... Im Umkreis 
der Kernkorperchen ist vielfach ein heller Hof, der von feinen 
Kornchen kugelschalenartig umgeben wird. Der Hof wird als 
Kernkorperchenhof beschrieben; er ist in sehr vielen Fallen 
sichtbar zu machen.” 

Davidoff (’ 89 ) observed in the egg of Distaplia a single 
large, spherical nucleolus, consisting of a homogeneous mass in 
which a few granules are imbedded. These nucleoli increase 
in size as follows : “ Sie werden grossere Partien des 

Reticulums in sich aufnehmen, sich mehr und mehr verdichten 
und demgemass sich immer deutlicher und deutlicher farben.” 
Subsequently, but antecedent to the production of the pole 
spindle, the nucleolus contracts, and its contour becomes 
irregular, often with regular branched processes: “Vielleicht, 
ja sogar wahrscheinlich, Werden sie dadurch hervorgerufen, 
dass der Nucleolus Fltissigkeit ausscheidet”; and the central 
portion of the nucleolus becomes lighter in color. Next, first 
the lighter portion, then the whole nucleolus, becomes filled 
with fine granules (“Chromatosomen ”). Then these chroma- 
tosomes collect and form in the center of the nucleolus 
a compact, granular body, in the middle of which is one 
especially large chromatosome, and the whole is surrounded by 
a membrane. And finally, other chromatosomes, not con¬ 
cerned in the formation of the central granular body, form a 
reticulum around it. Davidoff concludes “dass aus dem 
Nucleolus ein Kern mit Kernnetz, mit einem Nucleolus und 
Nucleolinus hervorgegangen ist. Wir konnen diesen Kern 
weder als Keimblaschen, noch als Nucleolus bezeichnen. Es 
ist eben ein neues Gebilde, dass wir einstweilen mit dem Namen 
Polkern belegen wollen”; out of this “Polkern” the first pole 
spindle is formed. 


MONTGOMERY. 


[Vol. XV. 


318 

Fol (’ 89 ), ovarian egg of Dentalium ; the nucleolus is at first 
absent, and single. In larger nuclei there are two apposed 
nucleoli (which disappear when the nuclear membrane has 
vanished). “Le nucleole presente d’abord deux parties dis- 
tinctes, dont l’une, plus volumineuse et moins fonc£e, entoure 
l’autre un peu comme un bonnet pose sur la tete. La partie 
foncee est spherique ; elle retient l’h^matoxyline ou le carmin 
alunique avec une nuance rougeatre ou vineuse. Sa texture 
est compacte. L’autre partie est formde des corpuscules plus 
clairs [vacuoles] et d’un r^seau plus fonce ; elle prend les colo¬ 
rants que nous venons de nommer avec une teinte violacee tirant 
sur le bleu. . . . Lorsque l’ovule approche de l’epoque ou la 
vdsicule germinative va se dissoudre, les deux nucleoles, au lieu 
de s’emboiter, sont simplement accoles, et le nucleole clair s’est 
accru beaucoup plus que l’autre.” 

Hermann (’ 89 a) investigated the spermatogenesis of the 
mouse. The “ Spermatoblastkern ” (nucleus of a v. Ebner’s 
cell) possesses one nucleolus, which is made up of two parts, 
“ einen von Safranin sehr intensiv gefarbten, und einen unge- 
farbt bleibenden Bestandtheil. Letzterer tritt stets in Form 
einer einfachen Kugel auf, die chromatische Substanz aber 
besteht entweder aus zwei kleinen, leuchtend roth tingirten, an 
zwei gegeniiberstehenden Polen der farblosen Kugel liegenden 
Kiigelchen, oder das chromatische Element stellt eine einzige, 
in diesem Falle grossere Kugel dar, die dem ungefarbten 
Bestandtheile des Nucleolus sich innig anschmiegt. Im 
ersteren Falle erscheint dann das ganze Kernkorperchen als 
ein annahernd spindelformiges Element, im anderen als eine 
Doppelkugel, und ist in beiden Fallen die Langsaxe des Nucleo¬ 
lus stets in dem grossten Durchmesser des Zellkerns einge- 
stellt.” The nucleoli of the spermatogonia are sometimes 
biscuit-shaped. Those of the spermatids are at first multiple 
in number, but later they unite to form a biscuit-shaped one. 
Still later, by the formation of the spermatozoon out of the 
spermatid, the two parts of this nucleolus wander apart, 
“ dabei aber noch durch eine chromatische Briicke mit einander 
in Verbindung stehen.” He observed in the follicle cells of 
the testicle of Salamandra “neben kleinen Nucleolen einen 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES . 319 

grosseren, . . . der vollkommen die gleichen Strukturverhalt- 
nisse zeigt, wie sie oben von dem Kernkorperchen der Sperma- 
toblastkerne der Maus beschrieben wurden und wie dies fur 
den Frosch von Sanfelice angegeben wird.” 

Hermann (’89b), testicles of immature white mice : the nuclei 
of the follicle cells contain compound nucleoli, similar to those 
of the cells of v. Benda of Salamandra. 

Korschelt (’89) made observations on the germinal spots of 
j Epeira, Dolomedes , Phalangium , Spinther , and Ciona. In Epeira 
the nucleolus is at first a compact mass of granules “von stark 
lichtbrechenden Kornchen umlagert. . . . Ich will damit nicht 
sagen, dass eine direkte Aufnahme von [nutritiven] Kornchen 
stattfande, welche letzteren sich dann unmittelbdr zum Keim- 
fleck formirten, sondern mochte vielmehr glauben, dass die 
Substanz in fliissiger Gestalt aufgenommen und erst im Kern 
wieder geformt wird”; in later stages small vacuoles are fre¬ 
quently present in the nucleolus. In Dolomedes the nucleolus 
is at first homogeneous, it later contains vacuoles, and finally 
becomes simply a membrane surrounding a cavity. In Spin¬ 
ther there is a single large nucleolus with a vacuole. Korschelt 
draws the general conclusion : “ Ich muss es nach meinen 

Erfahrungen, . . . als zweifelos hinstellen, dass eine Auflosung 
der Nucleolarsubstanz stattfindet. Die Erklarung dieser 
Erscheinung fand ich darin, dass die Nucleolarsubstanz in 
und vielleicht ausserhalb des Kerns zur Verwendung gebracht 
werden sollte.” 

Lukjanow (’89) describes the nucleoli ( “ plasmosomata ” ) of 
the germinal vesicle and cleavage nuclei; they disappear during 
mitosis. 

Platner (’89a), Malpighian tubule cells of Dytiscus , fixation in 
Kleinenberg’s fluid : there are one or several nucleoli, of irregu¬ 
lar form, and around each one usually “ein hellerer Hof, 
welcher aussen von einer Anzahl grosserer unregelmassiger 
Chromatinbrocken eingefasst wird.” The division of the nucle¬ 
oli introduces the amitosis of the nucleus : “ Der anfangs 
mehr runde Nucleolus zeigt eine Abplattung zur Scheibe, 
welche der umgebende Hof mitmacht. Zugleich tritt in der 
Richtung seiner kurzern Durchmesser eine Streifung an dem- 


320 


MONTGOMERY. 


[Vol. XV. 


selben auf, als wenn er aus einer Anzahl nebeneinander liegen- 
der schmaler Elemente zusammengesetzt ware. Weiterhin 
tritt eine Spaltung in der Richtung des langsten Durchmessers 
auf. . . . Die auf diese Weise entstehenden Tochterplatten 
zeigen an den einander zugewandten Seiten spitze Hervorrag- 
ungen, an den abgekehrten Flachen dagegen mehr abgerundete 
Erhabenheiten. Beide besitzen wieder eine langsgestreifte 
Struktur, als seien sie aus parallelen Stabchen zusammengefugt. 
. . . Den auseinanderweichenden Tochterplatten passt sich der 
helle, umgebende Hof an, der also in der Richtung dieser 
Bewegung sich verlangert.” 

Platner (’89b) contends, in opposition to the views of Ogata 
(’83) and others, that in the pancreas cells the nucleoli do not 
wander out of the nucleus. 

Platner’s (’89c) observations on the egg of Aulastomum shall 
be mentioned in the course of our observations on the egg 
of Piscicola. In accord with O. Schultze (’87) he finds in 
amphibian ova that the contents of the nucleus are composed 
only of “ Kernsaft und Keimkorperchen,” a portion of the latter 
forming the nuclear filament, the rest being extruded from the 
nucleus ; the true chromatin loops were not seen by him. 

Weismann and Ischikawa (’89) find in the ovarial winter ova 
of Leptodora one large nucleolus (rarely is a smaller one 
apposed to it), containing a large vacuole ; it wanders out of 
the nucleus and becomes the “ Nebenkern, Paranucleus,” 
which ultimately disappears, and corresponds to the nucleus 
alone of the paracopulation cell of the other Daphnids. In 
nearly ripe ova of Bythotrephes “ findet man . . . innerhalb des 
Keimblaschens und dem Nucleolus desselben ganz nahe einen 
scheibenformigen Korper, der sich wie der Nucleolus farbt. 
Etwas spater, wenn das Keimblaschen bereits an die Ober- 
flache des Eies gestiegen ist, liegt dieser Korper ausserhalb des 
Keimblaschens und ist in' einen Protoplasmahof eingebettet ”; 
then it rapidly disappears. 

Wheeler (’89), ovarial follicle cells of Blatta: there is a 
“nucleolus of unusual structure. The latter consists of an 
irregular mass, not stainable in carmine or methyl green, and is 
regarded as plastin by Carnoy. . . . The mass of plastin encloses 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


3 21 


a smaller mass of chromatin, or at least of a substance which 
does not differ in its reactions from the chromatin of the coiled 
filaments in the same nuclei.” This nucleolus divides first 
in mitosis. 


1890. 

Auerbach (’90) distinguished two kinds of chromatin sub¬ 
stance : “ erythrophile,” i.e., substances staining with eosin, 
fuchsine, aurantia, carmine, picrocarmine; and “ kyanophile,” 
substances staining with methyl green, aniline blue, haematoxy- 
lin. The nuclear reticulum is not the fundamental portion of 
the nucleus, but the nucleoli are its important elements. He 
finds “ dass in einer Grundsubstanz, die im frischen Zustande 
homophan, im geharteten . . . hochstens feinkornig erscheint, 
grossere, scharf begrenzte, isolirte, starker lichtbrechende und 
starker farbbare Korperchen, Nucleoli, von wechselnder,aber fur 
die verschiedenen Zellarten und Thierspecies typischer Anzahl 
eingebettet sind”; thus in the Batrachia most of the nuclei 
contain numerous nucleoli, and when they are particularly 
abundant the greater number are peripheral in position. 
There are two kinds of “ Kernkorperchen,” those which stain 
blue (or green) and those which stain red (or yellow) ; both 
kinds occur in most nuclei. In the giant nuclei of the gland 
cells from the skin of Urodelea are found ( 1 ) numerous small cyan- 
ophilic nucleoli, and ( 2 ) from one to fifteen (usually two to five) 
much larger, erythrophilic nucleoli, which sometimes contain vac¬ 
uoles. Embryonal nuclei contain only cyanophilic nucleoli, while 
in maturer nuclei erythrophilic nuclei become differentiated 
from the former. Thus in the blood corpuscles of frog larvae 
there is at first only one large nucleolus, which later differenti¬ 
ates into an inner erythrophilic and an outer cyanophilic por¬ 
tion. The peripheral layer next breaks up and divides into 
small cyanophilic nucleoli, while the central portion remains 
as a large erythrophilic nucleolus. Subsequently the smaller 
cyanophilic nucleoli (“ Nebenkiigelchen ”) may fuse together 
so as to produce six or eight larger cyanophilic nucleoli, each 
of which attains the size of the original “ Stamm-Nucleolus ”; 
at the conclusion of the larval period of the frog, the latter 


3 22 


MONTGOMERY. 


[Vol. XV. 


nucleolus entirely disappears, becoming dissolved in the nuclear 
sap. “ Die erythrophile Kernsubstanz ist iibrigens dem Pro¬ 
toplasma des Zellleibes offenbar ahnlicher als die kyanophile.” 

Burger (’ 90 ) made observations on the maturing ovum of 
various Nemerteans. Carinella: there is one large, spherical 
nucleolus. In the ripe egg of Cerebratulus marginatus “ in der 
Regel kann man zwei umfangreiche Keimflecke konstatiren, 
welche aus einer schwarzlich-grfinen kornigen Substanz zusam- 
mengesetzt sind, aber einen membranartig scharfen Kontour 
besitzen. Die beiden Keimflecke sind nicht von gleicher 
Grosse.” In the immature germinal vesicle of Drepanophorus: 
“ Dem wenig tingirten Binnenraum des Kernes durchflicht ein 
zartes Netzwerk feiner Faserchen ; peripher sind grobere dunk- 
lere Kornchen angeordnet ”; the ripe ovum of this Nemertean 
contains one finely granular, central nucleolus, in which are found 
“ kuglige, noch intensiver gefarbte Korperchen.” Prosadeno - 
porus janthinus: constituting the inner portion of the wall of 
the genital ducts are seen numerous cells, “ welche ganz wie 
in der Entwicklung im friihen Stadium stehen gebliebene 
Geschlechtsprodukte aussehen,” and each of these cells has 
one large nucleolus ; while in the ripe egg the “ Keimblaschen 
ist ausgezeichnet durch eine Menge kugliger Blaschen von fiber 
5 /x Durchmesser mit scharf kontourirter und stark gefarbter 
Peripheric.” 

Eimer (’90, cited by Mann, ’ 92 ), recalls his previous observa¬ 
tions (’73, ’78) in regard to the termination of nerve fibrils in 
the nucleolus ; he mentions further that such radiating fibers 
are also to be found in the nucleolus of the egg cell, such 
fibers serving at first as paths for nourishment, and later 
becoming nerve fibrils. 

Henking (’ 90 ), spermatogenesis of Pyrrhocoris: the single 
peripheral nucleolus of the first spermatocyte gradually 
becomes smaller in the prophase of division, and it is considered 
probable “dass er spaterhin eine Einschnfirung erfahrt.” 

O. Hertwig (’90), Ascaris megalocephala: in the spermato¬ 
cytes of the growth zone the nucleolus is usually flattened 
against the periphery of the nucleus, or it may be irregularly 
elongated, or in addition to it a “ Nebennucleolus ” may be also 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


323 


present ; from these differences in form he concludes that the 
nucleolus may be capable of amoeboid movements. Subse¬ 
quently it wanders towards the center of the nucleus, becomes 
larger and more spherical. When the chromatin has assumed 
the characteristic radial distribution, before the first maturation 
division, the nucleolus passes again towards the periphery, and 
there becomes gradually smaller, partly by fragmentation, and 
so gradually disappears. 

Holl (’90) found one spherical nucleolus in ova of the newly 
hatched chick: “ Da das Kernkorperchen so auffallend verschie- 
den vom Kernnetze und Kernsafte hinsichtlich des Verhaltens 
zur Farbe sich zeigt, so muss es wohl aus einem anderen Stoffe 
bestehen als jene. Auch bei Salamandra y Rana , und Lacerta 
fand ich das Kernkorperchen immer sich verschieden halten 
von den anderen Theilen des Kernes.” The nucleolus is always 
situated excentrically at the upper pole of the nucleus. Towards 
the end of the spirem stage the nucleolus lies on the periphery 
of the chromatin, with which it stands in no close connection; 
it is no longer present in ova of 491 / 1 , diameter. 

Kastschenko (’ 90 ) investigated the maturation of the ova of 
Pristiurus , Scyllium, and Torpedo: there are numerous nucleoli, 
which attain a diameter of 1 6 /x, and all disappear at the 
spirem stage (in the prophase of the first pole spindle). Each 
nucleolus contains a large unstaining globule (but in his Fig. 1 , 
in several of the nucleoli, all of which had been stained with 
borax carmine, this globule is colored blue, while the peripheral 
portion of the nucleoli is red). 

Masius (’ 90 ): in the ovum of Asplanchna the nucleolus forms 
the greater part of the nucleus. In Lacimdaria it is at first as 
in the preceding genus, but at a later stage several much smaller 
nucleoli are found. 

Mellissinos and Nicolaides (’90), pancreas cells of Canis: The 
“Nebenkern ” is a plasmosome which has wandered out of the 
nucleus; this migration is caused by an injection of pilocarpin 
into the living gland. 

Sheldon (’ 90 ) found one germinal spot in Peripatus capensis, 
which disappears when the nucleus reaches the periphery of 
the egg. 


3 2 4 


MONTGOMERY. 


[Vol. XV. 


Smirnow (’ 90 ), sympathetic ganglion cells of Rana and Bnfo : 
a “ Kernkorperchenkreis ” is figured around the nucleoli of 
some of the cells. 


i8gi. 

Brauer (’91) studied the maturation of the ovum of Hydra. 
As a rule in the smaller eggs there is a single large nucleolus 
which occupies an excentric position within the nucleus ; in 
larger ova numerous small nucleoli arise, which gradually 
become grouped near the large one. “ Die Anzahl [der kleinen] 
wechselt, was zum Theil darin seinen Grund zu haben scheint, 
dass der grosse — selten sind zwei grosse vorhanden — wahr- 
scheinlich durch Aufnahme kleinerer wachst . . . zum Theil 
aber auch darin, dass in verschiedenen Keimblaschen die 
Masse der Nucleolen eine verschieden grosse ist, was mit der 
Ernahrung zusammenhangen mochte. . . . Sehr oft lag in der 
Nahe des grossen Nucleolus eine etwa halb so grosse blasse 
Kugel . . . moglich ware es, dass diese sich vom grossen Nucle¬ 
olus abgespalten hat, und den achromatischen Theil derselben 
vorstellt.” Just before the formation of the first pole spindle the 
large nucleolus breaks into fragments, which, together with the 
smaller nucleoli, wander towards the periphery of the nucleolus : 
“ Ein Theil scheint im Keimblaschen selbst aufgelost zu werden, 
ein Theil tritt unverandert nach dem Schwinden der Membran in 
das Eiprotoplasma liber.” Brauer contends that the nucleoli 
have no morphological significance in the maturation of the egg. 

Cuenot (’91), ovarial egg of Synapta inhaerens: “ la tache 
germinative primitive bourgeonne une quantite de petits nucle- 
oles secondaires, qui errent dans le protoplasma clair de la 
vesicule germinative; presque toujours la tache a un aspect 
mamelonne par suite de la formation de ces nucleoles.” 

Davenport (’9i) figures in the germinal vesicle of Plumatella 
a double nucleolus. 

Macallum (’9i), following Ogata (’83), distinguishes two kinds 
of nucleoli, namely, plasmosomata and karysomata. He finds 
the “ Nebenkerne ” of Nussbaum to be abnormal structures. 
In the pancreas cells of Amphibia an extrusion of plasmosomata 
occurs, but it is not a normal process, and the extruded portion 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 325 

does not become a “ Nebenkern ” (in opposition to the views 
of Ogata). In the eggs of Rana and Necturus the chromatin is 
“ principally collected in the form of nucleoli at the periphery,” 
but it is also contained in certain threads in the nucleus. He 
concludes from the study of the reactions of the substances 
to the indigo-carmine stain: “ the peripheral nucleoli generate 
a substance, therefore, which diffuses gradually through the 
nucleus, then into the cell protoplasm, the point in time of the 
latter occurrence corresponding with the formation of the yolk 
spherules. The mode of origin is through a process of deposi¬ 
tion from the nucleus of a substance allied to chromatin in the 
cytoplasm. ... I regard the yolk spherules as formed by the 
union of a derivative of the nuclear chromatin with a constituent 
of the cell protoplasm.” 

C. Schneider (’91) concludes that the true spherical nucleoli 
“ ebenso wie die Klumpen [of the chromatic network] aus 
[achromatischen] Gerlist und Chromatin bestehen und die 
Unterschiede beider nur morphologischer Natur sind.” In the 
testicle cells of Astacns the nucleoli are spherical, with “ eine 
deutliche Membran an und durch welche genau wie bei der 
Kernmembran [achromatische] Geriistfaden treten. . . . Der 
ganze Unterschied zwischen Nucleolus und Klumpen besteht 
also hier darin, dass um ersteren die Fasern zu einer Membran 
sich zusammenlegen . . . was man ringformig am Rande des 
Nucleolus wahrnimmt, ist sicher nicht die optische Wiedergabe 
einer Membran, sondern durch das Brechungsvermogen der 
Wandung des Nucleolus veranlasst.” The nucleolus in eggs of 
Echinodermata is homogeneous only in the final stages of its 
formation. Nucleoli are only metamorphosed portions of the 
true chromatin, and represent reserve masses of this substance : 
“ die Zusammenballung kann nur eine Befreiung der chroma- 
tischen Substanz von ihrer Arbeitsleistung bedeuten.” 

Wolters (’91) studied the sporulation of Monocystis: in the 
youngest individuals there is one nucleolus, which “ in seinem 
Innern sich starker tingirende chromatische Kugeln fiihrt.” In 
larger individuals the nucleolus consists of eight spheres, “ Diese 
Kugeln fiihren in ihrem Innern wieder Stabchen und Korner.” 
Just before the conjugation of two individuals this compound 


3 2 6 


MONTGOMERY. 


[Vol. XV. 


nucleolus breaks into a number of nucleoli of various sizes. 
After the copulation and encysting the nucleoli fuse together 
and gradually disappear (but I am unable to determine from 
his description whether the substance of the chromosomes is 
derived from the nucleoli). Shortly after the nuclei themselves 
copulate, the nucleoli reappear in them. In Clepsidrina blat- 
tarum there is a single primitive nucleolus, formed as in 
the preceding species ; later there are numerous smaller 
nucleoli, which have probably arisen by division from the 
primitive nucleolus. 

1892. 

Bannwarth (’ 92 ) figures a division of the nucleolus in leuco¬ 
cytes from the spleen of the cat. 

Born (’92) finds that in the Amphibian egg, in opposition 
to the observations of O. Schultze (’87), the chromatic “ Faden- 
knauel ” has no origin in the nucleoli, but is directly derived 
from the chromatin network of the “Urei.” 

Brauer (’92) made observations on the maturation and fecun¬ 
dation of the egg of Branchipus. Each germinal vesicle from 
the “ Wachsthumszone ” of the ovary has one large, slightly 
staining nucleolus, and near it a much smaller, deeply staining 
one. Each “ Nahrzelle,” however, contains numerous nucleoli, 
and its nuclear sap also stains deeply. When the chromosomes 
are being produced, the larger nucleolus of the egg cell gradu¬ 
ally ceases to stain, and it finally disappears. In the male 
pronucleus small nucleoli are present. 

Frenzel (’92) noticed in Carcinus moenas and in a species of 
Amphipod , in the ferment cells and “ Fettzellen ” of the hepa- 
topancreas, amitotic division of the nucleus, but no division of 
the nucleolus ; “sondern dass vielmehr an geeigneter Stelle 
des Tochterkernes noch vor der Abschniirung desselben ein 
ganz neuer Nucleolus entstehe, der alle Charaktere des ersten 
besitzt ” ; in this nuclear division one of the daughter-nuclei 
retains the whole original nucleolus. In similar cells of Idotea 
tricuspidata he found the nuclear division to be as in the pre¬ 
ceding species (but his Figures 8b, 10 , and especially 11 , would 
seem to represent stages of division of the nucleolus). 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


Hacker (’92a) studied the early development of Aequorea 
forskalea: in the ripe egg there is one spherical or kidney¬ 
shaped nucleolus, containing vacuoles. At the time of the 
first pole body mitosis the nucleolus does not accompany the 
nucleus, but remains behind in the cell, at the place previously 
occupied by the nucleus ; and from this time on he applies to 
it the name “ Metanucleolus.” It is to be observed in one of 
the cleavage cells until about the 32 -cell stage. “ Zur Zeit 
wenn sich dann in der schwarmenden Blastula die Zellen des 
hinteren Poles . . . zu differenziren beginnen, kann man in 
einzelnen von ihnen neben dem chromatischen Fadenknauel 
kleine nucleolenahnliche Korper beobachten, welche den nicht 
differenzirten Blastula-Elementen fehlen. Es ware denkbar, 
dass man es hier mit den Abkommlingen des Metanucleolus zu 
thun hat, ich vermag aber weder hierrfiber, noch fiber das 
weitere Schicksal dieser Gebilde etwas bestimmtes zu sagen.” 
Hacker assumes that what Metschnikoff (’86) supposed to be 
the “ Sperm-nucleus ” in Mitrocoma was in reality a Metanucle¬ 
olus ; and also that the “ Paracopulationszelle,” described by 
Weismann and Ishikawa (’89) in the winter egg of Daphnia , 
to have been also a nucleolus. 

Hacker in a second paper (’92b) studied the maturation of 
the ovum of Canthocamptus . In the smallest eggs the nucleolus 
is large and contains vacuoles. Later it becomes differentiated 
into a lighter central portion and a denser peripheral part con¬ 
taining small vacuoles. At this stage the nucleolus presents a 
concavity facing the chromatic spirem. Then “ aus dem Kern- 
korper tritt unter plotzlicher Verkleinerung desselben eine Masse 
aus, welche vermuthlich dem grossen, bis dahin in den meisten 
Kernkorpern kugligen Einschluss entspricht.” The nucleolus 
apparently disappears when the first pole spindle is perfected. 

Heidenhain (’92), cells of Salamandra: the nucleoli lie 
enclosed within the chromatin and linin network ; he was unable 
to decide whether each nucleolus has a particular chromatin 
envelope. The nucleolus has no processes, and “ nur die ihm 
auflagernde, von ihm selber stofflich differente Schicht ist mit 
dem Chromatin- und Lininfadengerfist kontinuierlich verbun- 
den. . . . Mir ist es wenig wahrscheinlich, dass die Substanz 


328 


MONTGOMERY. 


[Vol. XV. 


der Nukleolen etwas dem Chromatin ahnliches sei. Zwar sind 
sie durch einige Chromatinfarbstoffe stark farbbar, wie z. B. 
durch Safranan, allein auf eine andere Gruppe derartiger Farb- 
stoffe reagieren sie nicht, hierher gehort das Methylgriin.” 

O. Hertwig (’92) in his recent text-book materially changes 
some of the views expressed in his previous papers. The 
true nucleoli consist of “ Paranuclein ” (Pyrenin), and he 
uses the term “Nuclein” for chromatin. “Nuclein und Para¬ 
nuclein betrachte ich als die wesentlichen Substanzen des 
Kerns. . . . Beide scheinen mir in irgend welchen Bezie- 
hungen zu einander zu stehen.” Further, he distinguishes 
“ Keimflecke ” from “echte Nucleolen.” “Je nach dem Alter 
oder der Entwicklungsstufe einer Zelle kann der ruhende Kern 
. . . in der Zahl, Grosse und Beschaffenheit seiner 4 Nucle¬ 
olen ’ erhebliche Veranderungen erleiden.” 

Kostanecki (’92a) is preliminary to his ’92b. 

Kostanecki (’92b) studied mitoses “ in samtlichen embryonalen 
Zellen ” of Lepus , Cavia , and Equus , with especial regard to the 
central spindle; I quote this paper here, since the “ Central- 
spindelkorperchen ” may have some relation to nucleoli. “ Im 
Bereich dieser Centralspindel sieht man in diesem Stadium 
[Dyaster] in der Nahe der beiderseitigen Tochterfiguren der 
Chromosomen kleine Korperchen auftreten, die ich als “Cen- 
tralspindelkorperchen ” bezeichnet habe. Grosse und Zahl 
dieser Korperchen zeigen ganz betrachtliche Schwankungen. 
. . . Meist fand ich nun jederseits vier, fiinf oder sechs gros- 
sere Korperchen, . . . daneben aber immer noch eine grossere 
Anzahl kleinerer Kornchen. Diese Kornchen sowohl als auch 
die grosseren Kornchen standen in inniger Beziehung zu den 
Faden der Centralspindel.” These granules then wander from 
both sides towards the equator of the spindle, “ so dass sie . . . 
eine aquatoriale Kornchenplatte bilden. . . . Sobald die Ein- 
schniirung des Zellleibes bis zur Centralspindel vorgeschritten 
ist, werden die mehr peripher gelegenen Centralspindelfasern 
gerade im Aequator da, wo die Centralspindelkorperchen liegen, 
durchschnitten, und man sieht die Korperchen zugleich mit den 
verkiirzten und undeutlich werdenden Fasern sich wiederum 
polwarts begeben.” At each pole, then, these granules become 


No. 2 .] COMPARATIVE CYTOLOGICAL STUDIES. 


329 


so densely grouped that often only one or two large granules 
appear to be present. “ Mit der volligen Durchschniirung 
der Zellen wird schliesslich . . . der Zwischenkorper in zwei 
Theile getrennt, von denen jeder einer Tochterzelle angehort.” 
Similar in the main points is also this process in the Chick , 
Frog, Axolotl , Triton , and Salamandra : “ Wenn wir uns nun 

fragen, ob diese Vorgange bei tierischen Zellen mit Recht mit 
den Vorgangen der Zellplattenbildung bei den pflanzlichen 
Zellen homologisiert wurden, so kann ich diese Frage nur zum 
Teil bejahen ”; for two processes take place together, “namlich 
eine aquatoriale Differenzierung der Centralspindelfasern zum 
Zweck ihrer Halbierung und eine eigentliche Zellplattenbildung, 
aus der die Zellscheidewand hervorgeht. Von diesen beiden 
Prozessen ist der eine, namlich eine eigentliche Zellplatten¬ 
bildung zum Zweck der Scheidewandbildung, bei tierischen 
Zellen gar nicht vertreten, wodurch der zweite desto deutlicher 
und unverhiillter zu Tage tritt.” (Kostanecki mentions the 
following observations of previous authors on the occurrence 
of such a granular aequatorial plate in animals: Van Beneden, 
germs of Dicyemida ; Balbiani, epithelial cells of an Orthopterous 
larva; Fol, eggs of echinoderms and Cymbtilia ; Flemming, 
eggs of echinoderms ; Biitschli, egg of Nephelis; Mark, egg of 
Limax; van Gehuchten, egg of Ascaris megalocephala; Pre- 
nant, testicle cells of Scolopendra and Lithobius; Henking, 
similar cells of Pyrrhocoris ; numerous observations of Carnoy ; 
Van Beneden, ectoderm of vertebrate embryos ; Strasburger, 
cartilage cells of vertebrates ; Mayzel, corneal epithelium of 
Fr ingill a; Schleicher, cartilage cells of Batrachia; Carnoy, 
Triton ; Biitschli, embryonal blood corpuscles of chick ; Schott- 
lander, inflamed epithelium of the cornea of the frog.) 

Kraepelin (’92, cited by Braem, ’ 97 ) noticed in the Bryozoan 
egg a division of the nucleolus. 

Lonnberg (’92) studied the nucleoli of various ova and somatic 
cells. In the liver cells of Mytilus there are two “ Nebennu- 
cleoli ” and one “ Hauptnucleolus.” In the cells of the intestinal 
epithelium of Tellina a granule is sometimes found on the 
outer surface of the nucleus, which resembles a small nucleolus, 
and stains in the same manner. Doris , egg: “ eine starker 


330 


MONTGOMERY. 


[Vol. XV. 


sich farbende Kugel (meist auch eine oder mehrere kleine 
Vacuolen) in einer grosseren hineingesenkt war und so den 
Nucleolus darstellte.” Mytilus: “In den Einucleolen von 
Mytilus liegt oft eine (oder bisweilen zwei) grosse, blasse 
Kugeln in der Mitte oder ein wenig excentrisch, aber von der 
starker sich tingirenden Substanz vollstandig umschlossen; es 
ist schwer zu unterscheiden, ob es sich hier nur um Vacuolen 
handelt. ... Bei Aeolidiapapillosa [Ei] . . . zwei, ein wenig 
abgeplattete Kugeln die in einanden teilweise eingesenkt sind. 
Diese Kugeln sind aber hier beinahe gleich gross und die 
blasse ist in der gefarbten eingesenkt, bei Unio [nach Flem¬ 
ming] umgekehrt. ... In den jungen Keimzellen fand ich 
nur einen einfachen Nucleolus, und dieser farbte sich stark.” 
In the liver cells of Doris proximo, there are two nucleoli: 
“ Der eine von diesen ist ganz kugelrund und stark licht- 
brechend, glanzend; dieser, der sich auch intensiv tingiert, muss 
als eigentlicher Nucleolus aufgefasst werden. Der andere ist 
blasser und grosser, seine Gestalt ist bald rundlich, bald lang- 
lich, bobnenformig also mehr unregelmassig; diesen mochte ich 
als Nebennucleolus bezeichnenthe two stain differently; 
“Die Lage der beiden Kernkorperchen ist auch wechselnd, 
indem sie bald ganz neben einander liegen oder sogar der Nucle¬ 
olus im Nebennucleolus hineingesenkt, bald vollig getrennt 
sind. . . . Der Nebennucleolus, der immer scharf begrenzt 
ist, enthalt oft eine kleine Vacuole. Ein Paar Mai traf ich in 
demselben Kern zwei Nebennucleoli.” The latter are homoge¬ 
neous, with an outer clearer layer, while the “ Hauptnucleolus ” 
is granular. Lonnberg found similar nucleoli also in the liver 
cells of Polycera and Aeolidia. Liver cells of the “Krebs” 
(Astacus ?): “ Meist sieht man . . . einen blassen Korper, 

der sich schwach wie der Nebennucleolus bei den Nudibranchi- 
aten farbt, und daneben einen oder mehrere kleine Korper- 
chen, die sich intensiv tingiren und sich wie Nucleolen verhal- 
ten ; . . . bald liegt ein stark gefarbtes Kiigelchen an einem 
Pole des Nebennucleolus, bald eins an jedem Pole desselben und 
in wieder anderen Fallen schmiegen sich drei Nucleolkorper- 
chen dem Nebennucleolus an. Bisweilen treten Nebennucle- 
olen in zwei- oder dreifacher Zahl auf.” Lonnberg concludes 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 331 

that the “ Nebennucleoli ” may play a part in the acquisition 
of nourishment or may hold reserve nourishment. 

Marshall (’ 92 ) studied the sporulation of Gregarina blattarum 
v. Sieb. In the youngest individuals there is one large nucle¬ 
olus. In larger ones there are one large and two or three 
smaller nucleoli, or four or five smaller ones of equal size ; 
these now increase in size, accompanying the growth of the 
nucleus. He believes that the smaller nucleoli which are sub¬ 
sequently produced, arise in only one (as a rule) of the four or 
five original nucleoli: “ Im Innern dieses Formationsnucleolus 
erscheinen dann klare, runde Ballen von verschiedener Grosse, 
welche keine bestimmte Grosse haben. Sie sind in wechselnder 
Zahl vorhanden und etwas heller als die ubrige Masse des 
Nucleolus. Bei vielen Formationsnucleoli . . . waren alle 
Stadien der Entwickelung zu finden; kleine und grossere 
Ballen im Innern, und einige, die schon halb nach aussen 
getreten waren.” After leaving the “ Formationsnucleolus ” 
they stain like the latter, and become either irregularly or 
spirally grouped together. “Die Vermehrung dauert bis zum 
Beginn der Cystenbildung fort. . . . Am Anfang der Encys- 
tierung enthalt jeder Kern etwa 25-40 deutlich erkennbare 
Nucleoli, welche bald in dieser, bald in jener Art angeordnet 
sind. In beiden Fallen liegt der jetzt unregelmassig gestaltete 
Formationsnucleolus der von ihm ausgegangenen Gruppe ge- 
geniiber” ; the latter is smaller than heretofore, “doch zeigt 
er noch Ballen im Innern.” The smaller nucleoli increase 
in number, but now by repeated divisions of their own; the 
small granules resulting from these divisions are termed 
“ Chromatinkorner ” : “Jedes Chromatinkorn bildet nun eine 
Htille um sich, nachdem es sich vorher mit einer Schicht Plasma 
umgeben hat. Auf diese Weise vollzieht sich die Bildung der 
jungen Sporen. . .. Kurze Zeit, nachdem die Spore gebildet ist, 
nimmt dieses Chromatin-Korn die Gestalt einer 8 an und teilt 
sich in zwei Halften, die beide an die entgegengesetzten Seiten 
der Spore treten.” Later each of these divides into two, and 
each of the resulting four then divides into two, so that eight 
is the result; then one such “Chromatin-Korn” is allotted to 
each “ Keim ” (young Gregarine) and represents its nucleus. 


332 


MONTGOMERY. 


[Vol. XV. 


Ruckert (’ 92 ) studied the maturation of the eggs of Scyllium , 
Pristiurus , and Torpedo. In young germinal vesicles there 
are a few small nucleoli, most of them peripheral in 
position. In larger ova they have increased in number and 
size, and become grouped in a cluster at that part of the nucleus 
which is nearest the animal pole of the egg ; this cluster may 
occupy one-fourth of the whole space of the nucleus. Later, 
but still antecedent to the formation of the pole spindles, the 
nucleoli decrease in size and commence to stain very faintly. 
Ruckert considers the nucleolus of an egg cell as strictly com¬ 
parable to that of any somatic cell. From the fact that the 
nucleoli are largest, and color most intensely, at the same time 
that the chromosomes do, and simultaneously with the latter 
become gradually invisible later, he concludes : “dass es die 
Stoffwechselvorgange der Chromosomen sind, zu welchen die 
Nucleolen in direkter Beziehung stehen, sei es nun, dass sie 
notwendige Stoffe an die letzteren abgeben (vielleicht das Chro¬ 
matin, wie schon Flemming vermutete), oder dass sie Stoffe 
von ihnen aufnehmen, Oder endlich dass beides zugleich der 
Fall ist. . . . Spater freilich, wenn die. Chromosomen merklich 
an Substanz verlieren, wird man eher geneigt sein, die betref- 
fenden Nucleolen als Trager von Zerfallsprodukten der Chromo¬ 
somen anzusehen.” He also observed that the number of the 
nucleoli varies in different germinal vesicles of the same age, 
that a number may coalesce to form a larger one, and that a 
few wander out into the cytoplasm, where they become paler 
and finally vanish. 

Wiren (’ 92 ) found that the smallest germinal vesicles of Chae- 
toderma contain no nucleoli; in nuclei of about 15ft diameter a 
nucleolus appears for the first time, and consists of a dense 
mass of granules, which stain differently from the other nuclear 
granules. More than one nucleolus is never to be found. 

1893. 

Van Bambeke (’ 93 ) found one to five homogeneous nucleoli 
in the germinal vesicles of Scorpaena scrofa , and notes that in 
older eggs they do not stain as deeply with carmine as in 
younger ones. 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


333 

Bohmig (’93), Rhodope veranii: the single nucleolus in older 
eggs contains one or several vacuoles. 

Brauer (’93) investigated the spermatogenesis of Ascaris 
megalocephala: there is one homogeneous nucleolus in the 
spermatogonium, which becomes smaller in the spermatocyte 
and often evinces a large vacuole. The nucleolus is smaller 
than the centrosome (which is at this stage enclosed in the 
nucleus), and stains differently from the latter. 

Brooks (’93), Salpa: the single large nucleolus of the ovarian 
ovum “ is suspended near the center of the nucleus by a net¬ 
work of fine threads.” 

Fick (’93) studied the maturation of the egg of the Axolotl. 
In the germinal vesicle lies a group of nucleoli, which vary in 
size from 3 /x to i6/x ; some contain a single vacuole, and some 
stain more deeply than others. The greater number of them 
disappear at the time of the longitudinal division of the chro¬ 
mosomes, though a few may remain in the yolk for a certain 
time. “Bei den Nucleolen des Keimblaschens liegt es sehr 
nahe mit Strasburger und Pfitzner daran zu denken, dass sie 
vielleicht eine Art Reservestoffbehalter darstellen ” ; further, 
he holds that the nucleoli “ in einer allerdings noch nicht auf- 
geklarten Beziehung zu den Veranderungen des Chromatins 
stehen, da sie bei der Ausbildung der Chromosomen fur die 
erste Spindel vollstandig verschwinden.” 

Frenzel (’93a) studied the nucleoli of various Gregarines. 
In Gregarina statirae the single nucleolus, which he terms 
“ Morulit,” appears “ eigenthiimlich glanzend mit einem 
schwach gelblichen Schimmer und dabei an der Oberflache 
rauh und warzig-runzelig. ... In seinen Reaktionen verhalt 
er sich an alien Orten ahnlich wie Nuklein.” In G. bergi a 
single Morulit is present. In the embryo of Pyxinia crystalligera 
there is a single Morulit; in older individuals the nucleus 
“ enthalt mehrere helle, klare, glattrandige und lebhaft glan- 
zende Nucleoli ... die oft noch einen Fliissigkeitsraum im 
Innern bergen.” In Gregarina port uni, Callyntrochlamys , and 
Aggregataportunidarum there are several nucleoli in the nucleus. 

Frenzel (’93b) hepatopancreas cells of Astacus: in the fat 
and ferment cells a single nucleolus is present ; in the amitosis 


334 


MONTGOMERY. 


[Vol. XV. 


of the nuclei he concludes that the nucleolus divides (“nukleo- 
lare Kernhalbierung ”), since the nucleoli of the daughter-cells 
are of equal size. 

Hacker (’93a) divides the maturation stages of the ovarian 
eggs of Moina , Cyclops , and Si da, into two periods, “vondenen 
der erste gekennzeichnet ist durch die Anwesenheit eines 
einzigen ‘Nucleolus’ und durch die leichte Farbbarkeit des 
Fadenspirems (chromatische Stufe), der zweite durch die 
Anwesenheit mehrerer ‘ Nucleolen ’ und die Abneigung der 
chromatischen Substanz, die Mehrzahl der Farbungsmittel 
anzunehmen (achromatische Stufe).” In the first period 
(“ Wachstumsphase ”) there is one excentric, deeply staining 
nucleolus (“ Hauptnucleolus ”), which possesses a “ Hullmem- 
bran”; in the second period, in addition to the “Hauptnucle¬ 
olus” there are also one or two “ Nebennucleoli ” of greater 
size than the former, but staining less deeply, and somewhat 
irregular in form. Both kinds of nucleoli contain vacuoles. 
The “ Nebennucleolus . . . stellt sich vielfach als hohles 
Gebilde von ellipsoidischer Gestalt dar, dessen einem Pole der 
Hauptnucleolus kappenformig aufsitzt.” Only the outer shell 
of this nucleolus stains deeply. Subsequently the “ Haupt¬ 
nucleolus” grows gradually smaller and finally disappears; 
and at the same time the “Nebennucleolus” increases in size 
and becomes irregularly lobular in shape, and finally breaks 
into pieces. The nucleolar relations in Moina are as in Cyclops 
(just described). In Si da only a “ Hauptnucleolus ” is present, 
and this contains a large central and several smaller peripheral 
vacuoles. Hacker distinguishes the following types of ova 
with regard to their nucleolar structure : (i) Lamellibranchiate 
type , with one “ Hauptnucleolus ” and one or two “ Neben¬ 
nucleoli,” the latter larger and less chromatic than the former, 
but both frequently in close connection (Naja, Anodonta, 
Cyclops brevicornis ) ; ( 2 ) Echinoderni type , with one large 
“ Hauptnucleolus,” which increases in size, and only towards 
the close of the “ Keimblaschenstadium ” do a few smaller 
nucleoli appear (Toxopneustes, Sida crystallina , primiparous 
Cyclops strenuus and C. signatus ); ( 3 ) Vertebrate type , with 
several nucleoli varying in size, number, and form ( Rana and 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


numerous other Vertebrates , Sagitta , Moina , Cyclops brevicornis, 
multiparous C. strenuits). “Aus der obigen Zusammenstel- 
lung . . . geht . . . hervor, dass das unter der Bezeichnung 
* Nucleolus’ oder ‘Keimfleck’ im Eikern auftretende Gebilde 
hauptsachlich in zweierlei Gestalt auftritt : entweder stellt 
dasselbe einen in der Einzahl vorhandenen, stetig seine Grosse 
verandernden, formbestandigen Korper dar, oder aber finden 
sich als ‘ Nebennucleolen’ Blaschen oder Tropfchen von wech- 
selnder Zahl, Grosse und Gestalt vor.” The “ Hauptnucle- 
olus ” remains in the nucleus until just before the formation of 
the first pole spindle ; after that it either diminishes rapidly in 
size, or it passes out of the nucleus into the cytoplasm, where 
it remains for a time as a “ Metanucleolus the “ Hauptnucle- 
olus ” is phylogenetically derived from a “ Nebennucleolus,” 
and has developed into “einen membranumhullten, formbe¬ 
standigen und stetig durch Diomose wachsenden Organ.” 
“Es diirfte vielleicht zunachst die Thatsache heranzuziehen 
sein, dass ein Auftreten von ‘ Nebennucleolen ’ von wechselnder 
Zahl, Form und Grosse und von analoger chemischer Reak- 
tion auch in den ruhenden Furchungskernen der betreffenden 
Thierformen festzustellen ist, und dass diese 4 Nebennucleolen ’ 
hier nicht mit einem als Hauptnucleolus anzusprechenden 
Korper vergesellschaftet sind. Nebennucleolen treten folglich 
auch da im ruhenden Kerne auf, wo kein Zellenwachsthum 
stattfindet.” Hacker considers that the “ Nebennucleoli ” are 
not drops of a nutritive fluid, but “Abspaltungsprodukte oder 
Sekretstoffe der chromatischen Substanz. Diese Auffassung 
findet vor allem in der Thatsache eine Stiitze, dass die Neben¬ 
nucleolen, z. B. bei Moina und Cyclops strenuus (mehrgebarend), 
im Lauf der Wachsthumsphase stetig an Grosse und Massigkeit 
zunehmen und dass sie das Maximum ihrer Entwickelung erst 
in dem Moment erreichen, wenn bereits die Vierergruppen zur 
Ausbildung gelangt sind, wenn also von einem Wachsthum der 
chromatischen Substanz kaum mehr die Rede sein kann.” 

Hacker (’93b) : a preliminary contribution to the following 
paper. 

Hacker (’93c) found in the germinal vesicle of Echinus micro - 
tuberculatus , in addition to the “ Hauptnucleolus,” a few small 


33 ^ 


MONTGOMERY ,i 


[Vol. XV. 


globules which stain in the same manner as, and are probably 
homologous to, the “Nebennucleoli” of other animals; the 
“ Hauptnucleolus ” increases in size by the absorption of these 
latter. “Der Hauptnucleolus des Echiniden-Keimblaschens 
ist . . . ein pulsirendes Organulum, in welchem periodisch eine 
grosse Hauptvacuole sich durch Zusammenfluss kleinerer 
Vacuolen bildet, urn dann wieder langsam abzunehmen. . . . 
Was die Dauer der Perioden anbelangt, so wurden solche von 
vier bis zu solchen von acht Stunden beobachtet ”; the central 
vacuole at the time of its maximum size passes from the cen¬ 
ter to the periphery of the nucleolus : “ Die Centralvacuole 
tritt also in Beziehung zur aussersten Wandschicht des Haupt¬ 
nucleolus, anscheinend um ihren Inhalt mit . . . den Kernsaft 
in Kommunikation zu bringen.” Accordingly, the “ Haupt¬ 
nucleolus ” may be “ als ein osmotisches System betrachtet 
werden, in welchem die feste Substanz (Rindensubstanz) nach 
zwei Seiten hin, einerseits mit dem Kernsaft, anderseits mit 
den Vacuolen, in diosmirender Verbindung steht. Sobald 
jedoch ein Korper nach zwei Seiten diosmirt, so ist eine An- 
haufung in demselben nur durch das Eingehen einer neuen 
Verbindung moglich [Pfeffer (’9l)]. Es folgt schon hieraus, 
dass die aus dem Kernsaft aufgenommene P'liissigkeit in der 
Nucleolarsubstanz nicht nur eine Verdichtung, sondern auch 
eine weitere chemische Umsetzung erfahren muss.” The fluid 
vacuoles of the “ Hauptnucleolus ” represent an excretion 
which in Echinus is periodic, while in the Copepoda “ im Laufe 
der Eireife wachst unter Mitwirkung der Rindenvacuole die 
Centralvacuole langsam heran, nimmt allmahlich eine excen- 
trische Lage an und entleert sodann kurz vor der Bildung der 
Richtungskorper ihren Inhalt nach aussen.” He compares the 
vacuole of the “ Hauptnucleolus ” to the pulsating vacuole of 
the Infusoria: “ so wurden die Centralvacuole des Hauptnucleo¬ 
lus mit der eigentlichen pulsirenden Vacuole des Protozoen- 
korpers, die Rindenvacuolen des Hauptnucleolus mit den 
Bildungsvacuolen zu vergleichen sein.” From a study of the 
pole-body mitoses he concludes : “dass der Hauptnucleolus 
wahrend der Auflosung der Keimblaschenwandung zunachst 
noch in seiner urspriinglichen Grosse erhalten bleibt und sich 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


337 


von dem Kernplasma langsam zu trennen beginnt.” He could 
not exactly determine how long the nucleolus remains in the egg 
after that, but considers the fact important, “ dass der Haupt- 
nucleolus zur Zeit der Umbildung der Keimblaschensubstanz 
ohne bemerkbare Volumverminderung fort besteht ”; it is at 
this time (after the disappearance of the nuclear membrane), 
to use his terminology, a “ Metanucleolus.” 

Henneguy (’93) studied principally the genesis and occur¬ 
rence of the yolk nuclei, “corps vitellin de Balbiani,” in the 
ovarian egg of various vertebrates. This body is absent in the 
ova of the Rabbity Bitchy Mole y RhinolophiiSy Cow f Antelope , 
Baboon; and in Lizards , Galeus y Raja, and ScyIlium. In the 
rat it consists of a peripheral clearer portion and a central 
denser core ; it stains with eosin, safranin, and haematoxylin, 
but not with methyl green or gentian violet. In the bat it 
encloses a spherical corpuscle. It is also present in the chicken. 
Though absent in Bnfo and Triton y it is found in Rana tempo- 
rariay where it is much the same as in birds, enclosing a more 
deeply staining portion. In the trout the corpuscle of Balbiani is 
as in the rat, but larger (twenty). Sygnathus: the very young 
egg contains a nucleus “ renfermant un r6seau chromatique bien 
developpe ” (his Fig. 20 would show three small nucleoli enclosed 
in this “ reseau ”); in older eggs the nuclear membrane is lined 
by a large number of nucleoli, and “ le centre du noyau est 
occupe par une petite masse finement granuleuse et teintee en 
rose par la safranine, tandisque le reste du contenu demeure 
incolore. Le protoplasma ovulaire est dgalement faiblement 
colore et renferme un corpuscule arrondi, refringent comme les 
taches germinatives et retenant la safranine avec la meme 
intensite que ces derni&res ” ; this corpuscle at the time of its 
first appearance is flattened against the outer surface of 
the nuclear membrane. Subsequently this intravitelline body 
becomes elliptical in form, with its long axis parallel to the surface 
of the egg : “ il est de plus au contact immddiat par son bord 
externe avec une amas arrondi, constitue par une substance 
fondamentale d’apparence homogene, mais remplie de granula¬ 
tions tres colordes. A un stade plus avancd tout le corps 
refringent s’est transforme en un amas let qu’on Tobserve dans 


338 


MONTGOMERY, 


[Vol. XV. 


la plupart des ovules des Poissons ” ; in this manner it develops 
into a Balbianian corpuscle, and later breaks into small granules. 
In no eggs of any of the species studied are more than one 
of these corpuscles present; and it always arises during the 
maturation period of the ovum, before fecundation. “ C’est 
tres probablement une partie de la tache germinative, ou une 
tache germinative entiere, qui sort la vesicule [germinative] 
pour penetrer dans le vitellus. . . . C’est un organe ancestral 
qui, avec les elements nucleolaires de la vesicule germinative, 
correspond au macronucleus des Infusoires, le micronucleus 
etant represente par le reseau chromatique, prenant seul part 
aux phenomenes de fecondation.” 

Heuscher (’93) noticed in the ovum of Proneomenia either 
one nucleolus, or two of different size which were usually 
separated from each other. 

Holl (’93) studied the maturation of the ovum of the mouse. 
“Die Faden [Chromatin] zeigen eine innige Verbindung mit 
dem Kernkorperchen derart, als ware dasselbe ein Centrum, 
von welchem die Faden des Netzwerkes auslaufen.” The 
nucleolus is not homogeneous, but contains granules (“ Schroen’- 
sche Korner ”) to the number of twenty ; these gradually become 
stained during the growth of the nucleus, until the whole nu¬ 
cleus becomes evenly stained. “ Im weiteren Verlaufe der 
Entwickelung treten die Schroen’schen Korner aus dem Kern¬ 
korperchen heraus und gelangen als chromatische Ballen in 
das Kernnetz, wo sie sich mit den Faden desselben verbinden. 
. . . Endlich wird das Kernkorperchen von seinem Inhalte ganz 
frei; es bleibt nur die Kernkorperchenmembrane iibrig, und im 
Kernraume liegen zerstreut eine grossere Anzahl der chroma- 
tischen Ballen. Dieselben sind anfangs klein und schwach 
gefarbt, wachsen auf 2/jl heran und farben sich immer besser. 
. . . Die chromatischen Ballen wandern aus dem Kerne aus, 
und das iibrige [Fadennetzwerk] riickt als ‘ Kernrest ’ ganz an 
die Oberflache der Eizelle. Die chromatischen Ballen liegen 
in 6 Gruppen von je 4 neben einander, und jeder Ballen wandelt 
sich in eine dicke, kurze Schleife urn,” i.e., a chromosome of the 
“ Richtungsspindel.” 

Jordan (’93) studied the development of the ovum of the 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


339 


newt. He thinks “that certain deutogenic substances are 
formed in the [germinal] vesicle, perhaps through the agency 
of the nucleoli, and are then sent forth to share in the building 
up of the cell,” i.e., of the yolk particles. “The nucleoli in 
the young egg appear arranged along the chromatin threads, 
and possibly originate from the thread substance.” Later they 
lose this connection, grow larger, and assume a peripheral 
position within the nucleus. There is apparently no division 
of the nucleoli; they “ attain their maximum size shortly 
before their centripetal movement.” Having arrived at the 
periphery of the nucleus, the nucleoli commence to stain less 
deeply, their contours become uneven, and they then wander 
back to the center of the nucleus, where they disintegrate. 
He does not agree with O. Schultze (’87) that the nucleolar 
particles build up the chromosomes. 

Kaiser (’93) found in the egg of Echinorhynchus bipennis one 
large, spherical, peripherally situated nucleolus. It disappears 
before the pole spindle is produced. 

Lustig and Galeotti (’93), mentioned by Lardowsky (’94), con¬ 
sider that the centrosome does not proceed from the nucleolus. 

Mertens (’93), ovum of Homo: two or three nucleoli are present, 
consisting of a central clearer and a peripheral darker portion; 
it is probable that several smaller ones may fuse together to 
form a larger one ; they are at first in intimate connection with 
the chromatin filaments, but later lose this connection and gradu¬ 
ally cease to stain with safranin. The Balbianian corpuscle 
is an extruded nucleolus : “c’est alors aussi que nous nous 
^tondrons quelque peu sur l’expulsion des parties chromatiques 
du noyaux, expulsion qui parait affecter les memes charact&res 
chez les oiseaux et les mammiferes ”; eliminated nucleoli 
(“ grains chromatiques ”) as well as attraction spheres have been 
described as Balbianian corpuscles. Ovum of Pica: in young 
ovules there is one nucleolus which arises as follows : at one 
point in the nucleus the reticulum concentrates itself, and 
here a certain number of the filaments fuse together, thus 
producing the nucleolus. The chromatin is at first irregularly 
arranged in the nucleolus, but “ finit par etre egalement dense 
dans toutes les parties de la tache germinative,” and subse- 


340 


MONTGOMERY. 


[Vol. XV. 


quently accumulates on its surface. “ Le nucleole devenu 
independant [from the chromatin reticulum] est expulse : les 
chromosomes s’ecartent pour lui livrer passage. II n’est pas 
rare d’en rencontrer qui, arrives a la peripherie, sont coiffes par 
un filament nucl&nien. . . . Le filament se rompt bientot et le 
nucleole est libre.” The presence of a vacuole in the nucle¬ 
olus is explained by the assumption that the chromatin wanders 
to the periphery of the nucleolus, thereby leaving a clear 
space at the center of the latter. (Safranin the only stain 
employed.) 

Minchin (’93) states that the single nucleolus of Gregarina 
irregularis “ consists of a darkly stained ground substance con¬ 
taining an immense number of clear vacuoles of all sizes. One 
of the vacuoles is much larger than the others, and being 
excentrically placed, constitutes the clear spot seen in the 
thick sections.” The nucleolus of G. holothuriae has a similar 
structure. 

Pizon (’93), ova of Botryllida: a single large nucleolus con¬ 
taining several vacuoles. 

Repiachoff (’93) figures a large vacuole in the single nucleolus 
of the ovarial cells of a pelagic, acoelic Rhabdocoele (species 
undetermined). 

From Rhumbler’s contribution (’93) to the morphology of the 
nucleoli, or “ Binnenkorper,” the following extracts are impor¬ 
tant : “ Mir scheint es . . . noch keineswegs sicher, ob die 
Nucleolen der Gewebszellen und die Nucleolen der Keimzellen 
bezw. vieler Protozoen (vielleicht ausgenommen die Ciliaten 
und Suctorien) analoge Gebilde sind ; obgleich auch das Gegen- 
theil wegen des ahnlichen Verhaltens der beiderlei Nucleolen- 
arten wahrend der Mitose sehr zweifelhaft bleiben muss.” In 
Saccamina sphaerica there are from i to 300 nucleoli: “ ahnlich 
wechselnd wie ihre Zahl ist ihre Grosse, ihr Lichtbrechungs- 
vermogen und ihre Gestalt.” The largest of them “zeigen 
meist eine, durch starkeres Lichtbrechungsvermogen ausge- 
zeichnete Innenmasse, in welche kleinere, noch starker bre- 
chende und oft von der Kugelgestalt abweichende unregelmas- 
sige Korperchen eingelagert sind, und eine dunklere, weniger 
lichtbrechende Aussenmasse, die in gleichmassiger Dicke wie 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 341 

eine feste Membran um die Innenmasse herum gelagert ist ”; 
this latter portion also stains more intensely with eosin. 
Rhumbler concludes that the “ Binnenkorper . . . durch Zu- 
sammenfliessen anfanglich leicht fliissiger, dann zahfliissiger 
und schliesslich erstarrender Massen entstanden sind. Ich 
nehme an, dass die Binnenkorpersubstanz an alien oder auch 
nur an bestimmten Stellen (das Letztere da, wo eine fixirte 
Nucleolenzahl Regel ist)* des Kernplasmas zuerst in Gestalt 
kleinster, erstarrender Tropfchen abgeschieden wird, die auf 
verschiedenen Stadien ihrer Erstarrung an einander treffen,” 
this deduction being based in part on an observation of A. 
Schneider (’75). He explains why the nucleoli are not evenly 
distributed in the nucleus, on the ground “dass die einzelnen 
Tropfchen jedenfalls nicht an alien Stellen des Kernraumes zu 
genau derselben Zeit entstehen.” The nucleoli probably repre¬ 
sent “ Reservestoffe,” which are consumed in the later growth 
of the nucleus, and since in Saccamina they decrease in size 
as the amount of the chromatin increases, it is probable “ dass 
die Nucleolensubstanz [die sehr verschieden sein kann] in irgend 
welcher Beziehung zum Chromatin steht.” Further, he holds 
that the nucleolar substance is produced in the nucleus, “ und 
dann erst erzeugt wird, wenn sie in kleinen Tropfchen auf- 
tritt.” But it is not yet possible to decide whether the nucle¬ 
oli of the Metazoa also arise in this manner, and hence the 
use of the general term “ Binnenkorper ” instead of the more 
specific one “ Nucleolus.” That amoeboid movements of nucle¬ 
oli have been noticed is not contradictory to his theory, since 
changes of form would be caused by the processes of fusion, 
or these motions might denote “ Auflosungsvorgange ”: “ Die 
Auflosung der Binnenkorper muss nach unserer Annahme von 
zwei, im Kernsaft enthaltenen, sich gegen die Binnenkorper 
kontrar verhaltenden Substanzen, auf eine Ueberschreitung 
des angestrebten Mischungsoptimums von Seiten der losenden 
Substanz zuriickgefuhrt werden. . . . Der Verschmelzungsvor- 
gang ist schon von mehreren Forschern erschlossen oder ver- 
muthet worden — neu diirfte nur die Annahme einer allmahli- 
chen oder auch rascheren Erstarrung der ursprunglich flussigen 
Binnenkorpersubstanz sein.” Rhumbler concludes that the 


342 


MONTGOMERY. 


[Vol. XV. 


“ Binnenkorper ” are not organs, since they show no fixed 
organic structure, but represent accumulations of various sub¬ 
stances. There is more nucleolar substance, “ Reservestoff,” 
accumulated in the nucleus before mitosis than is necessary 
for it, so that after a mitosis some always remains to serve for 
the production of daughter-nucleoli (this being an explanation 
for the reappearance of nucleoli after mitosis). 

Stauffacher (’93), maturation of the egg of Cyclas: the 
“ Urei ” contains a single large nucleolus ; later one or two 
“ Nebennucleoli ” also appear in the nucleus. When the ovum 
has so increased in size that it adheres to the wall of the ovary 
only by a narrow thread of cytoplasm, two nucleoli are present, 
which are of unequal size but are in close contact with each 
other; in one case the nucleolus was trilobular. After borax- 
carmine staining, the smaller one appeared more refractive and 
deeply stained than the larger. Subsequently the two became 
separated, and both vanished before the formation of the first 
pole spindle. 

Strasburger’s paper (’93) presents a general discussion of 
certain problems of mitosis in animals and plants; his 
remarks on the aequatorial plate are apropos here. He believes 
that the “Kornchen” found by Kostanecki (’92) in the equator 
of the central spindle are similar to, and comparable with, 
structures found by himself in the mitoses of plants, and are 
masses of nucleolar substance (these bodies being termed 
“ Centralspindelkorperchen ” by Kostanecki, “ Zwischenkor- 
per” by Flemming, and “ Zwischenkiigelchen ” by O. Hertwig, 
’92). “ Vergegenwartige ich mir nun das, was ich seinerzeit bei 

der Bildung pflanzlicher Zellplatten beobachtet habe \Histol. 
Beitr ., vol. i, p. 161 ], namlich das Fortschreiten jener tingir- 
baren Substanz, die ihrem Auftreten und ihren Tinctionen 
nach nur als Nucleolarsubstanz gelten konnte, zwischen den 
Verbindungsfaden bis zum Aequator, so muss in mir die Vor- 
stellung erwachen, dass es sich in der von v. Kostanecki 
geschilderten Erscheinung um einen entsprechenden Vorgang 
handle. . . . Mit den durchschnittenen [achromatischen] Ver¬ 
bindungsfaden . . . wanderte dann auch die Substanz der 
halbirten Zwischenkorper nach den Zellkernen zuriick, ahnlich, 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES . 


343 


wie wir das fur die unverbrauchte Nucleolarsubstanz bei Pflan- 
zen angeben konnten. ... Bei Pflanzen treten die Elemente 
der Zellplatten als Anschwellungen der Verbindungsfaden im 
Aequator der Zelle auf. Diese Anschwellungen bilden sich 
dort erst, wenn jene tingirbare Substanz . . . den Aequator 
erreicht. Diese Substanz wird in geloster Form zwischen den 
Verbindungsfaden dorthin befordert. Aus den verschmolzenen 
Elementen der Zellplatte geht die Scheidewand hervor. . . . 
Man konnte denken, dass in tierischen Zellen ein mittlerer 
Teil der * Zwischenkorper ’ in eine losliche Substanz sich 
verwandle und so die Halbierung der Zwischenkorper und 
damit auch der Verbindungsfaden bewerkstelligte.” 

Ver Eecke (’93), pancreas cells of Rana and Canis: he 
distinguishes one “nucleole nucldinien ” and several “nucleoles 
eosinophiles,” or plasmosomes, the latter being the larger. 
When the cell enters on its functional activity “ le plasmosome 
unique devient plus volumineux; il n’est pas rare d’en trouver 
plusieurs dans un seul noyau; ils se rapprochent de la mem¬ 
brane nucleaire, la soul&vent, la perforent et se placent en 
definitive a cote du noyau pour former un noyau accessoire. 
D’ordinaire le plasmosome dans sa migration est accompagne 
de petits karyosomes qui lui forment parfois une veritable 
couronne”; the mother-nucleus subsequently degenerates. 
Against the opinion of Platner (that the supposed migration 
of the nucleoli is artificially produced) “ il suffit de faire remar- 
quer que la migration ne s’observe pas ou tr&s rarement a 
l’etat de repos pour ne se manifester dans tout son eclat 
qu’au ddbut de l’activite secretaire.” In the cytoplasm the 
nucleolus and its attendant karyosomes gradually change into 
a nucleus. 

Wasielevsky (’93) found the “ Urgeschlechtszelle ” of Ascaras 
with one or two nucleoli. While in the resting state of the 
nucleus only one nucleolus is present, two are regularly seen 
in the spirem stage, and these he believes have originated by 
division of the primitive one. He noticed no difference in size 
or stain between these nucleoli and the centrosomes, and hence 
concludes that the latter are identical with, or have some 
genetic relation to, the former. 


344 


MONTGOMERY. 


[Vol. XV. 


1894 . 

Blochmann (’94) gives a preliminary account of the results 
of the observations of Keuten (’95). 

Born (’94) investigated the maturation of the ovum of Triton. 
In the “ Urei ” are one or several large, spherical nucleoli. In 
the second stadium of the maturation (production and degenera¬ 
tion of a “ Chromatinfadenknauel ”) there are at first ten nucle¬ 
oli, then they become more numerous, increase in size, and 
lie close to the nuclear membrane. In the third stadium (eggs 
of from 200ft to 350ft in diameter) the nucleoli increase still 
more in size. In the fourth stadium (eggs measuring from 350ft 
to 800ft, first appearance of yolk in the cytoplasm) most of the 
nucleoli lie in the peripheral “ Karyohyaloplasma,” only a few 
pale ones being in the center of the nucleus (this part of the 
nucleus he terms “ Centralkorper ”). At the commencement 
of this stage the nucleoli increase, at its conclusion decrease, 
in number, and “wahrend der ganzen Periode steigt die Zahl 
der verkleinerten und abgeblassten Nucleolen im Centralkor¬ 
per,” only a few of these pale ones being situated at the 
periphery of the nucleus. Thus while at the beginning of this 
period the nucleoli attain their maximum size, at its end most 
of them wander towards the center of the germinal vesicle, 
become smaller, and lose their staining power. Fifth stadium 
(the nucleus passes to the periphery of the egg): the nucleoli 
decrease still further in size, and continue to wander to the 
center of the nucleus ; some of the larger ones contain vacuoles, 
and for the first time appear granular ; the smaller, lightly 
staining nucleoli are division products of the larger ones. At 
the commencement of the sixth stadium (formation of the first 
pole spindle) all the nucleoli lie in irregular rows around 
the “ Centralkorper,” stain quite intensely, and are regularly 
vacuolated ; the few in the midst of the “ Centralkorper ” are 
smaller and stain more faintly ; when the nucleus has decreased 
still further in size, all the nucleoli vanish at once. Born con¬ 
cludes as follows : “ Eine sichere Herleitung der peripheren 
Nucleolen von den Nucleolen des Ureies, bin ich freilich nicht 
im Stande zu geben. . . . Die Nucleolen stehen in Beziehung 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


345 


zum individuellen Zellleben, nicht zur Fortpflanzung; denn beim 
Beginn der Mitose verschwinden sie, um nach Beendigung der- 
selben — im Ruhestadium des Kerns — wieder aufzutreten.” 
He notes that their peripheral position is “ Eine Lage, die fur 
eine Wirkung auf den Zellleib die denkbar giinstige ist.” 

Brauer (’ 94 ), Actinosphaeriiim : in the cyst of the second order 
(“ Ruhecyste ”) there is a nuclear reticulum consisting of 
chromatin granules imbedded in a linin network, and usually 
numerous nucleoli of irregular form, arranged either in rows or 
circles. Probably the nucleoli take no part in the formation 
of the chromosomes, and are equivalent to those of metazoan 
cells ; they disappear in the prophase of mitosis. 

Bunting (’ 94 ) found in the eggs of Hydractinia and Podoco- 
ryne a single large nucleolus, containing one central vacuole of 
large size. 

Flemming’s (’ 94 ) “ Referat ” includes some of the more 
recent papers on nucleoli. 

Foot (’ 94 ), egg of Allolobophorci: during the first maturation 
division the nucleoli are distributed in the cytoplasm. Each 
pronucleus contains from one to seven nucleoli: “ the nucle¬ 
oli persist during the cleavage spindle, but how much later I 
am unable at present to state.” 

Hodge (’94), nerve cells of Rana stimulated by the electric 
current : amoeboid movements of the nucleolus were noticed; 
“ it was possible to make out granules in the nucleolus which 
moved slowly about and in several instances were seen to be 
extruded into the nucleus”; and in cells which had not been 
stimulated, but simply fixed in osmic acid and stained with 
safranin, “ the granules were stained brighter red than 
the body of the nucleolus, and several were, found partially 
extruded.” 

Lavdowsky (’ 94 ) studied nuclei from the epidermis of the 
fins of Amphibian larvae, as well as various tissues of plants. 
The nucleolus consists of : (i)an outer, thick “ Pyrenin-Chro- 
matinschale ”; (2) an enclosed vacuole ; and in the latter ( 3 ) 
the “Nucleololus ” (“das noch in Entwickelung begriffene 
Centrosoma”). The animal nucleolus varies from a spherical 
to an angular or star shape. In the resting nucleus the chro- 


346 


MONTGOMERY. 


[Vol. XV. 


matin and pyrenin shells are the largest, since “die Bestand- 
theile noch nicht fur die Karyokinese verbraucht sind.” The 
centrosomes “sind wahrscheinlich Teile von Kernkorperchen 
und wandern zur Zeit der Karyokinese von den Kernelementen 
aus” (these centrosomes are spherical or oval, homogeneous 
and compact, and stain very slightly). He concludes “ dass die 
Kernkorper nicht zu jeder Zeit des Zellenlebens persistieren, 
dass ihr Verschwinden wahrend der Karyokinese keinem Zweifel 
unterliegt und dass dies in innigem Zusammenhang mit dem 
Erscheinen des Centrosoma steht.” The nucleoli divide amito- 
tically (not seen in life, however) into very small pieces, which 
“ scheinen in das Geriistnetz eingeschaltet und verwandeln sich 
in den Vorbereitungsstadien der Karyokinese in Chromatin- 
faden”; other “Kernkorper” pass out of the nucleus, at the 
points where its membrane is broken. The nucleoli are not 
sufficient for supplying the whole mass of chromatin necessary 
for the mitosis ; “es muss also eine andere Quelle der Chromatin- 
entwicklung da sein und hauptsachlich im Eidotter und in den 
pflanzlichen Samen muss man die Quelle aufsuchen. . . . 
Durch nichts unterscheiden sich die Chromosomen von den 
zerteilten Dotterkornchen und den geteilten Nucleolen. Alle 
diese Gebilde . . . konnen somit als 1 Kariosomen * betrachtet 
werden.” 

Metzner (’94), cells in the testicle of Salamandra: he con¬ 
cludes “ dass die Nucleolen in keinem Stadium der Mitose 
fehlen, obwohl sie von sehr verschiedener Grosse sind.” In 
resting nuclei the smaller nucleoli stain entirely with gentian 
violet (after Flemming’s triple stain), the larger ones with 
safranin except for a blue-stained peripheral zone. Smaller 
nucleoli are budded off from the surface of the larger ones, and 
the “ Leitkorper ” (granules which serve to attach the chromo¬ 
somes to the spindle fibers) resemble such buds in stain and 
size ; “ es ist mir vorerst nicht moglich zu entscheiden, ob diese 
Leitkorper von dem Nucleolus stammen, doch ist es wahr¬ 
scheinlich, dass gerade an ihm sich die ersten, den Kern- und 
Zelltheilungsprocess einleitenden Vorgange abspielen. Denn 
an den Zellen mit ziemlich gleichmassiger Vertheilung der 
Chromatingranula und geringer Anzahl der Nucleolen kann 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 347 


man immer schon den Vorgang der Ausstossung kleinster 
Kugelchen beobachten.” In mitosis the nucleoli wander into 
the cytoplasm, where the larger of them disappear, while the 
smaller persist; “ dass aber diese Nucleolen in den Tochterkern 
einwandern, ist nicht sehr wahrscheinlich, denn es liegen in den 
Tochterzellkernen nur die gelosten Leitkorper. . . . Vielleicht 
persistiren nur einige von ihnen in der jungen Zelle und zwar 
als Nucleolen. . . . Eine ‘Vermischung von Nucleinsubstanz’ 
kann ich ... an meinen Praparaten . . . fiir Chromatingranula- 
strange nicht annehmen, denn die augenscheinlich von den 
Nucleolen stammenden Leitkorperchen adhariren nur den 
Segmenten und erfiillen ihre . . . Function als Anheftungs- 
punkte der Spindelfibrillen; sie losen sich intakt in den 
Tochterknaueln wieder ab. Dass sie noch andere Functionen 
ausiiben (als Nucleolen) ist wahrscheinlich, doch nicht ganz 
sicher. . . . An den Nucleolen treten die ersten Erscheinungen 
der Zelltheilung auf. Sie lassen aus sich eine Menge kleiner 
Kugelchen hervorgehen, die z. Th. aus dem Kerne in das Pro¬ 
toplasma wandern, z. Th. aber als Leitkorperchen iiber den 
Kern sich vertheilen und so wohl den Anstoss geben zur 
Strangbildung der Chromatingranula. Dem Nucleolus fiele also 
fiir die Fortpflanzung der Zelle eine wichtige Function zu.” 

Murbach (’94) considers it probable that the “ Kapselkeim ” 
of the nettle capsules of hydroids is derived from one of the 
two nucleoli of the parent cell, in accordance with his view that 
the capsule is of nuclear origin. 

Purcell (’94) describes the nucleolus of the retinula cells of 
Acantholophus as structurally “wabig.” 

Reinke (’94) found in the cells of the spleen of the mouse 
one oval or elongate nucleolus ; during the prophase of mitosis 
this divides into three or four smaller ones, while at the end of 
mitosis each daughter-nucleus has a single nucleolus. 

Riickert (’94) studied the maturation of the ovum in three 
species of Copepoda. Cyclops strenuus (his species he assumes 
is not identical with the “C. strenuus” of Hacker) : in the 
“ Wachsthumszone ” of the ovary there is one large, sometimes 
also two smaller nucleoli, which stain with haematoxylin as does 
the chromatin, and together represent the “ Hauptnucleolus ” 


34 § 


MONTGOMERY. 


[Vol. XV. 


of Hacker. The single “ Nebennucleolus ” appears a little later, 
and is regular in its occurrence, both in females with egg 
sacks {“ mehrgebarend,” after Hacker) and in those without 
egg sacks (“erstgebarend,” according to Hacker); Hacker found 
the “ Nebennucleolus ” only in the ova of the former category 
of females. It is paler and much larger than the several 
“ Hauptnucleoli,” and has a more central position within the 
germinal vesicle, while the latter are usually peripheral. When 
the “ Hauptnucleoli ” have disappeared the “ Nebennucleolus ” 
increases in size and thereby at first assumes a mulberry shape, 
or is produced into long processes (though at the start it was 
spherical). “ Wahrend er anfanglich ein kompaktes Gefiige 
besitzt, lockert er sich spater auf. Schon friihzeitig sieht man 
in seinem Innern einen lichteren Raum, und spater entrollt er 
sich zu knauelartig gewundenen Ziigen, die ein sehr wechselndes 
Ansehen bieten, sehr haufig bilden sie eine einzige, ziemlich 
einfach verschlungene Figur, ein Achtertour, ein S u. a., neben 
der jedoch noch ein oder ein paar kleinere kugelige Stiicke im 
Keimblaschen liegen konnen. ... Er ist . . . nicht einheitlich 
gebaut und homogen, wie ihn Hacker abbildet, sondern zusam- 
mengesetzt aus rundlichen Anschwellungen, die in einer Reihe 
hinter einander liegen, stellenweise getrennt durch schwach 
gefarbte, schmalere Zwischenstiicke. Man konnte daher das 
Ganze als eine Kette von Kugeln bezeichnen. ... In etwas 
spateren Stadien verlieren diese Bildungen an Farbbarkeit, 
erscheinen aber zunachst immer in sehr wechselnder Form. 
Man trifft entweder einen mehr kompakten Substanzhaufen 
oder meistens eine Anzahl durch das Keimblaschen zerstreuter 
Stiicke. . . . Haufig sieht man ein vielfach verschlungenes, 
sehr unregelmassig angeordnetes Fadensystem. ... Es ist 
schwierig zu entscheiden, ob die beschriebenen, sehr wechsel- 
vollen Bilder der Ausdruck nur fur verschiedene Functions- 
zustande des Nucleolus sind, oder fur einzelne, zum schliess- 
lichen Zerfall fiihrende Entwicklungsstufen they disappear 
before the true maturation processes commence. In Heterocope 
robusta and Diaptomus gracilis there is a single large, vacuolated 
nucleolus; it disappears when the chromatin has arranged 
itself into “ Vierergruppen.” 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


349 


Schaudinn (’94) finds in Amoeba crystalligera a large nucle¬ 
olus, with “wabiger Struktur”; in the mitosis it divides into 
two equal parts. 

Watasd (’94), in the course of his theoretical deductions as to 
the structure of the cell, concludes in regard to the nucleolus: 
“ The nucleolus is not a permanent body in the nucleus. It 
may exist at one stage of the cell, and may disappear at the 
next. The micro-chemical reaction of the nucleolus is entirely 
different from that of the chromosome. It appears probable 
that three or more different bodies are included under the name 
of nucleolus. Indeed, one sees no reason why the inside of the 
nuclear membrane may not be used as a depository for some 
solid products of cell metabolism. . . . And thus some of the 
bodies included under the generic name of nucleolus may 
belong to the group of metaplasm.” 

H. V. Wilson (’94), Tedanione foetida: the youngest germinal 
vesicle contains a single, centrally placed nucleolus. Later 
there are two nucleoli, “ which are invariably placed on opposite 
sides of the nucleus and adhere to the inner surface of the 
nuclear membrane. In eggs which have reached the adult size 
it is the rule to find either one nucleolus peripherally placed, 
... or the nucleus contains no nucleolus at all. It sometimes 
happens that an egg of full size is found with two nucleoli, but 
this is rare. From this evidence it would seem that the two 
nucleoli present in the developing egg are lost, one after the 
other, at the time when the egg reaches its full size. As to 
how the first of the two is lost, I have no evidence, but the 
second nucleolus may often be seen lying just outside of the 
nucleus in the yolk, . . . showing that it has been extruded 
from the nucleus.” What Fiedler (’ 88 ) described as polar 
bodies in Spongilla are probably extruded nucleoli. In the egg 
of Hircinia acuia the nucleolar changes are as in Tedanione. 

1895 . 

Balbiani (’95), reviewed by v. Erlanger in Zool. Centralbl ., 
1895 , macronucleus of Spirochona : the nucleolus of the authors 
arises in a vacuole of the chromatin, and is formed by the 
separation of microsomes which fuse together to form one 


350 


MONTGOMERY. 


[Vol. XV. 


or two nucleoli. The nucleolus then wanders through the 
chromatin to take position in the center of the achromatic 
substance ; it combines the qualities of a true nucleolus with 
those of a centrosome. There is thus no fundamental differ¬ 
ence between a nucleolus and a centrosome ; when it remains 
in the nucleus it has the value of the former, when in the 
cytoplasm it has the significance of a centrosome. 

Bohmig (’95) noticed in the ovarial eggs of Haplodiscus that 
the nucleolus is at first small and homogeneous, while later it 
becomes larger, and one or more vacuoles appear in it. 

Bremer (’95a), blood cells of Testudo and Chelydra: there is 
normally one paranuclear corpuscle to a cell ; “ seiner Natur 
nach ist das Paranuclearkorperchen ein vom Innern des Kernes 
in das Diskoplasma [Cytoplasma] ausgewanderter Nucleolus 
oder vielleicht ein Nucleolusfragment, umgeben von einer dem 
Kerne entnommenen Hiillsubstanz. . . . Seine Grosse, die 
Schwierigkeit der Farbung und seine Lage sprechen fur den 
nucleolaren Charakter.” In a second paper (’95b) he identifies 
this corpuscle with a centrosome, and states : “ Hertwigs 
Vermuthung, dass ein Zusammenhang des Centrosoms mit dem 
Nucleolus existire, wird durch meine Beobachtungen wahr- 
scheinlicher gemacht.” 

In Burger’s monograph (’95) of the Nemerteans the following 
statement in regard to the structure of the germinal vesicle is 
of interest: “ Im Keimblaschen findet man ausser den intensiv 
farbbaren Korperchen, den Nucleolen, von denen meist zwei, 
ein grosseres und ein kleineres, vorhanden sind, ein Netzwerk 
feiner Faden, in welche sehr feine Kugelchen aufgehangt sind.” 

Coe (’95), ova of Cerebratulus lacteus: “ as the ovum 

increases in size its nucleus develops into the germinal vesicle 
which has many germinal spots, of which one or two are much 
larger than the others.” In the mature ovum the nucleus 
“often contains a highly refractive germinal spot one-third as 
large as the vesicle itself.” 

Cunningham (’95), ovarial eggs of fishes : in the youngest 
ova there is a single large nucleolus, in older ova a number of 
peripheral ones ; the latter are produced in part by a division 
of* the primitive nucleolus, in part by an increase in size of 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 351 

" minute nucleolar granules ” which were present in early stages. 
In contradiction to the view of Scharff (’88) he finds that no 
nucleoli wander out of the nucleus to form yolk globules. 

Delage (’95) opposes the view that the nucleoli and the 
centrosomes are genetically related (as against the theory of 
Julin (’93 b) and Wasielevsky). 

Galeotti (’95), embryonal cells of Triton and Spelerpes (fixa¬ 
tion in Hermann’s fluid with chloride of palladium substituted 
for chloride of platinum ; stained for five minutes in sat. sol. 
acid fuchsine in aniline water at 6o° C., then stained in 
sol. methylen green in equal parts of water and alcohol for three 
or four minutes): “ Auf diese Weise erhalt man roth gefarbt 
die Kornchen des Cytoplasma und alle Elemente des Kerns mit 
Ausnahme des Nucleolus . . . ; gelblichgriin erscheint der 
protoplasmatische Grund der Zelle und lebhaft grim die baso- 
philen Granulationen.” In the pancreas cells of Spelerpes the 
green-stained nucleolus passes out of the nucleus and persists 
as “ Nebenkern,” which in the cytoplasm seems to increase by 
continued division ; and from this he concludes “ dass der 
Nucleolus ein endonucleares Arbeitsprodukt des Kernes ist, 
bestimmt aus der Kernmembran auszutreten und im Cytoplasma 
so umgeandert zu werden, dass er in Secretionsprodukte umge- 
wandelt wird.” 

Hacker (’95) first describes the nucleolar relations in the eggs 
of Canthocamptns , and then gives expression to general views, 
based on his numerous previous observations, in regard to the 
nature of nucleoli. Canthocamptns staphylinus : in the smallest 
eggs there is one large nucleolus, which increases in size, but not 
to same relative extent as does the nucleus itself; subsequently 
vacuoles arise in it, one of which becomes a “ Hauptvacuole ”; 
smaller “ Kernkorper ” appear first when the chromatin elements 
commence to thicken ; “ wenn endlich die Kernsubstanz auf 
das Minimum ihres Volumens zusammengedrangt ist, so fehlt 
in der Regel jede Spur von nucleolarer Substanz.” Then 
follows his general conclusions in regard to the physiology and 
structure of the nucleoli : “ Die Nucleolen sind nach meiner 

Ansicht im allgemeinen als nicht strukturirte Gebilde aufzu- 
fassen. . . . Sie stellen als solche ... ein Abspaltungsprodukt, 


352 


MONTGOMERY. 


[Vol. XV. 


welches wahrend der vegetativen Thatigkeit der Zelle und des 
Kerns in oder an den chromatischen Elementen zur Abschei- 
dung gelangt und zu Beginn der Mitose aus dem Kernraum 
entfernt wird. Wie bei alien organischen Wachsthums- und 
Umbildungsprocessen, so wiirden . . . Sekret-Substanzen zur 
Abspaltung kommen, welche in Form eines Hauptnucleolus 
oder mehrerer Nebennucleolen auftreten. . . . Die Griinde, 
welche theils fur die Auffassung der Nucleolen als nicht 
organisirter Stoffwechselprodukte sprechen, theils speciell 
darauf hinweisen, dass es im Kern entstandene und dem Kern 
verlassende secretartige Stoffe sind,” are the following : (i) 
“ Die bedeutende Entfaltung der nucleolaren Substanz in den 
Kernen solcher Zellen, fur welche eine intensive vegetative 
Thatigkeit angenommen werden muss (Keim-Mutterzellen, 
Driisenzellen, Ganglienzellen, Wimperzellen), wiirde zum min- 
desten dafiir sprechen, dass die Nucleolarsubstanz ein Stoff- 
wechselprodukt darstellt, dessen Erzeugung in einem gewissen 
Abhangigkeitsverhaltniss zur Intensitat der vegetativen Leis- 
tungen von Kern und Zelle steht.” He cites numerous cases 
to show that all germ cells with little yolk and with usually 
adequal cleavage have a large “ Hauptnucleolus ” (sponges, 
Hydromedusae, Siphonophora, Acalephae, Ctenophora , Echino- 
dermata , Copepoda , Tomopteris ) ; while all large ova with a 
considerable amount of yolk and with discoidal or superficial 
cleavage have numerous “ Nebennucleoli ” (most Insecta , many 
Crustacea , lower Vertebrata). He explains the time of the 
appearance of the “ Nebennucleoli ” in the egg of Canthocamptus 
in this way : “ Zur Erklarung dieser Erscheinung ist anzuneh- 
men, dass irgend welche die ganze Eizelle betreffenden Ver- 
anderungen physiologischer Natur, die um diese Zeit eintreten, 
die weitere Apposition der neu sich bildenden Nucleolarsub¬ 
stanz an den Hauptnucleolus verhindern und das Auftreten 
mehrerer Verdichtungscentren hervorrufen, welche haufig nicht 
mehr das Farbungsvermogen des urspriinglichen Hauptnucleo¬ 
lus erlangen . . . vom rein morphologischen Standpunkt aus 
darf man aber wohl mit diesen in den Endstadien der Eibildung 
auftretenden Bildern jeden intermediaren Keimblaschentypus 
vergleichen, welcher sich im Lamellibranchiaten-Ei vorfindet.” 


No. 2 .] COMPARATIVE CYTOLOGICAL STUDIES. 


353 


He notes that “ die Bildung nucleolarer Substanz auch unab- 
hangig vom Zellwachsthum in erheblichem Masse stattfinden 
kann. Bekanntlich treten namlich auch in den zur Copulation 
sich anschickenden Geschlechtskernen Nucleolen auf, welche 
nicht selten betrachtliche Dimensionen annehmen, und dasselbe 
gilt fur die Kerne der friiheren Furchungsstadien. Hier ist 
von einem Zellwachsthum nicht die Rede.” Accordingly, he 
concludes: (i) “dass die Menge der nucleolaren Substanz in 
einem direkten Verhaltniss steht zur Intensitat der Wechselbe- 
ziehungen zwischen Kern und Zelle”; ( 2 ) “hier mochte ich 
nur wiederholen, dass ich aus den verschiedenen Bildern eine 
Entstehung der Substanz der Nucleolen an oder in den Chro- 
matinschleifen und die Moglichkeit einer Verschmelzung der- 
selben ableiten und mich so entschieden gegen die Auffassung 
aussprechen mochte, dass die Kernkorper aus dem Zellplasma 
in den Kern hereingelangen und hier in die Bildung des Chro¬ 
matins eingehen, sowie im allgemeinen dagegen, dass die 
kleinen durch Theilung der grosseren entstehen ”; ( 3 ) he 
brings a few observations to show “ dass der Kern die nucleo- 
lare Substanz an das Zellplasma abgiebt, dass es sich also hier 
wohl kaum um Stoffe handelt, welche als Nahrmaterial dem 
Chromatin zugefiihrt werden, sondern um solche, die wahrend 
der Veranderungen des letzteren zur Abspaltung und dann zur 
Ausscheidung aus dem Kerne kommen. . . . Ich denke . . . , 
dass sie [die vorhergehenden Erorterungen] in ihrer Gesamt- 
heit sehr wohl eine Stiitze fur die Kernsekret-Theorie bilden 
konnen.” Finally, Hacker gives his own explanation of the 
maturation stages of Triton , based on the description of Born 
(’94), and, comparing the changes here with those observed by 
himself in the maturation of Canthocamptus , generalizes the two 
as follows : 1 . Stadium (growth of the germinal vesicle), “ Ab- 
scheidung einer dunkel tingirbaren Nucleolarsubstanz 2 . 
Stadium, “ Verdichtung der chromatischen Substanz und Con- 
centrierung in die Kernmitte. Beginn der Auflosungsvorgange. 
Der neu sich bildende Nachschub an nucleolarer Substanz 
erlangt . . . nicht mehrdas urspriingliche Farbungsvermogen ”; 
3 . Stadium, “ Grossenreduktion des Keimblaschens : Die chro- 
matische Figur liegt unmittelbar im Zellplasma.” 


354 


MONTGOMERY. 


[Vol. XV. 


Held (’95) finds that in the ganglion cells of vertebrates, 
when stained with erythrosin followed by methylen blue, the 
nucleolus stains blue and the “ Nebennucleoli ” violet. 

Herrick (’95) found that the nucleolus of Homarus contains 
one large and several smaller vacuoles ; the gravitation of the 
nucleolus in the caryolymph, i.e., its movement to the lower 
side of the nucleus, may be post-mortem phenomena (at least I 
learned as much from Dr. Herrick during a brief conversation). 

Keuten (’95) investigated the nuclear division of Euglena 
viridis. In the nucleus there is an elongate body, the “Nucle- 
olo-Centrosoma,” which stains more intensely than any other 
portion of the nucleus. At the commencement of mitosis it 
elongates, “ und wahrend die Segmente [Chromosomen] bisher 
eine annahernd senkrechte Richtung zur Oberflache des Nucle- 
olo-Centrosomas eingenommen hatten, bilden sie jetzt einen 
spitzen Winkel mit demselben,” and gradually come to lie 
parallel to it. At this time the middle piece of the “ Nucleolo- 
Centrosoma” stains more lightly than its ends, so that these 
latter parts are sharply demarcated from it (with the stain of 
Heidenhain, namely, Bordeaux R. followed by haematoxylin). 
“In der folgenden Phase riicken die parallel zum Nucleolo- 
Centrosoma gelagerten Chromosomen von beiden Polenden her 
nach dem Aquator zu, so dass die Enden des Nucleolo-Cen- 
trosomes nunmehr frei in die Kernhohle hineinragen, wahrend 
die Chromosomen als breite aquatoriale Zone das Mittelstiickdes 
Nucleolo-Centrosomes umgeben.” Next, the nucleus assumes 
the form of a rotation ellipse, in the short axis of which the 
“Nucleolo-Centrosoma” lies. After the longitudinal splitting 
of the chromosomes, from three to five vacuoles appear in each 
end of the “ Nucleolo-Centrosoma ” ; then the latter structure 
elongates and breaks into two parts, while at the same time the 
long axis of the nucleus gradually changes so as to coincide 
with the long axis of the “ Nucleolo-Centrosoma,” and part of 
the chromosomes become grouped around the one end, the 
remainder around the other end, of the “Nucleolo-Centrosoma.” 
Keuten believes his “Nucleolo-Centrosoma” to be comparable 
to the nucleolus of Amoeba crystalligera (Schaudinn), to the 
“ Centralspindel ” in Diatomea (Lauterborn), and to the centro- 


No. 2.] COMPARATIVE CVTOLOGICAL STUDIES. 


355 


some plus central spindle of Ascaris megalocephala; it is 
probably an important mechanical factor in the mitosis. 

Korschelt (’95) finds that in the amitosis of the intestinal 
cells of Ophryotrocha puerilis the “ Kernkorper ” divides into 
two. Ovarial and cleavage stages of the same annelid : the 
“ Kernkorper ” in the cleavage cells arises as “ eine Anhaufung 
von Chromatin, die sich zu einer Kugel abrundet. In ihr taucht 
bald eine polygonale Felderung als Ausdruck einer schon ganz 
friih beginnenden wabigen Struktur des Kernkorpers auf.” The 
“ Kernkorper ” increases in size rapidly, attaining its maximum 
size and staining intensity when the chromatin filament for 
the next mitosis becomes well marked. From this time on 
“ beginnt sein allmahlicher Verfall ” ; it stains less intensely, 
owing to the walls of its meshes becoming thinner ; the regu¬ 
larity of the latter becomes lost, and granules appear within 
and between them, while at the same time the “ Kernplasma ” 
[“ Kernsaft ”] stains more deeply: “Wahrend vorher das 
Kernplasma hell und der Nucleolus dunkel gefarbt erschien, 
hebt sich jetzt umgekehrt der helle Kernkorper von dem 
dunklen Kernplasma ab. . . . Immerhin halte ich es fur 
moglich und sogar fur wahrscheinlich, dass zu dieser Zeit ein 
Austausch zwischen dem Kernsaft und der geformten Substanz 
des Kernes stattfindet, bei welchem vielleicht ein Theil des 
vorher im Kernkorper niedergelegten Chromatins dem Kern- 
faden beigefiigt wird.” Similar nucleolar changes take place 
in the male and female pronuclei, antecedent to the stage of 
the first cleavage spindle ; in the male pronucleus “ man sieht 
. . . bei dem aus dem Kopf des Samenfadens sich herausbil- 
denden Spermakern im Geriistwerk den Nucleolus auftauchen.” 
The younger germinal vesicles contain one deeply staining, 
homogeneous “Kernkorper”; later vacuoles arise in it, so that 
it eventually evinces an alveolar structure ; the time when the 
nucleolus disappears is quite variable, thus it may sometimes 
remain when the chromatin filament is perfected: “ Dieser kann 
iibrigens auch noch vorhanden sein, wenn die vier Kernschleifen 
bereits gebildet sind. Das letztere Verhalten mochte man 
entschieden so denken, dass die Substanz des Kernkorpers von 
keinerlei Bedeutung fur die Ausbildung der chromatischen 


356 


MONTGOMERY. 


[Vol. XV. 


Substanz ist. Das oben eingehend besprochene Verhalten der 
Embryonalkerne liess dagegen eine ganz andere Auffassung zu, 
obwohl es auch bei diesen allerdings abnormer Weise vorkommt, 
dass neben den bereits gebildeten Chromosomen (sogar in der 
angelegten Spindel) der Kernkorper noch vorhanden ist. . . . 
Was die erwahnten Verschiedenheiten des Verhaltens der 
Nucleolen in dem Ei- und Embryonalzellen betrifft, so liessen 
sich diese vielleicht durch die recht verschiedenartige Ausbil- 
dung und Funktion der Kerne in den beiderlei Zellen erklaren.” 

Lauterborn (’95a), nuclear division of Ceratium hirundinella: 
from one to four oval nucleoli are present and are frequently 
apposed to the nuclear membrane. One nucleolus is still 
present in the spirem stage (the mitosis advances no further 
than this); but he was unable to decide whether this nucleolus 
divides into two. 

Lauterborn (’95b), Multicilia : each nucleus contains a rela¬ 
tively large nucleolus, which frequently shows a “netzig- 
wabige” structure. 

Macallum (’95) concludes that less iron is contained in the 
nucleolus than in the chromatin, as is shown by its lighter stain 
with haematoxylin. Nucleoli “are always attached to the 
chromatin network, and sometimes there appears about them a 
membrane derived from, and continuous with, the fibrils with 
which they are connected.” In a nucleus of a gland cell from 
the kidney or liver of Necturus “ which is passing into the 
mitotic phase, the nucleolar body disappears, apparently by 
solution into the chromatin threads, for in the nucleus of a renal 
cell, in which the meridional disposition of the chromatin 
filaments obtained preparatory to the formation of the loops, I 
saw, attached to one of the filaments and partly embraced by 
its substance, what appeared to be the remains of such a body.” 
The nucleoli of the amphibian ovum are derived from the 
chromatin of the nuclear reticulum. In support of his previous 
observations (’91) he adds, “ that the iron in the cytoplasm of 
the ovum makes its appearance only after the solution of the 
peripheral nucleoli commences.” In plant cells ( Erythronium) 
there are at least three kinds of nucleoli : the first stain 
intensely with eosin ; the second are composed of chromatin ; 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


and the third kind, which occur in the embryo sac, “ are not 
present in the mitotic nucleus, but in the retrogressive stage 
[metaphase] they appear on the course of the filaments as 
spherical elements enclosing one or more refracting corpuscles 
and containing but a small amount of iron, which, however, in 
later stages ... is more abundant. These nucleoli are eventu¬ 
ally formed chiefly of chromatin, and in stained preparations 
appear to contain nearly all the chromatin of the nucleus. 
When mitosis again commences the filament forms at their 
expense, the increase in size of the filament keeping pace, 
apparently, with the decrease in the quantity of chromatin 
which the nucleoli contain. Finally, before their disappearance, 
when they contain but a minimal quantity of iron, they take 
the eosin stain deeply. All these forms of nucleoli take up 
safranin from solutions as readily as do the chromatin elements 
in the same nuclei, and they hold the stain as tenaciously when 
they are washed with alcohol. They are in this respect differ¬ 
ent from the eosinophilous nucleoli in the animal cell, which 
appear to be unrepresented in the vegetable cell.” In Spirogyra 
and Corallorhiza “ the greater portion of the chromatin in each 
nucleus forms a single large spherical element unconnected with 
the chromatin network.” He corroborates Leydig’s view of 
the structure of the chromatin loops in the nuclei of the salivary 
glands of Chironomus; the nucleolus is often vacuolar and 
amoeboid, and may be transversed several times by the chro¬ 
matin loop ; “ the presence of granules and vacuoles . . . 
appears to indicate that it is physically active, which cannot be 
postulated of the vast majority of the nucleoli of Vertebrate 
cells.” In Euglena the nucleolus stains deeply with eosin 
(except after fixation in picric acid), but does not stain with 
safranin ; it is “ intermediate in composition between the nucle¬ 
olus of higher animal cells and the chromatin of the nuclear 
reticulum.” 

Mead (’95), egg of Chaetopterus: “in the second cleavage, as 
in the first, the nucleoli are dropped out into the cytoplasm in 
the equatorial plane.” 

Montgomery (’95) described the various arrangements of the 
nucleoli (“ Chromatinmassen ”) in the ova of Stichostemma 


MONTGOMERY. 


[Vol. XV. 


353 

eilhardi. “Was diese Chromatinmassen chemisch darstellen, 
ist mir vollig unklar: vielleicht sind sie als von dem Dotter 
aufgenommene Nahrsubstanzen zu betrachten, oder vielleicht 
stellen sie Konglomerate mehrerer Kernsubstanzen dar.” (In 
my present paper I have no corrections to make to these 
previous observations, but add only fuller descriptions of the 
genesis of these nucleoli.) 

Moore (’95), spermatogenesis of Selachii: the resting nuclei 
of the first spermatogenetic period contain each a single large 
nucleolus, which disappears in the following mitosis. In the 
subsequent resting stage the nucleolus reappears, and also there 
appears a smaller “ secondary nucleolus ” surrounded by a 
vacuole. The larger one then “ takes a position, generally, but 
not always, in line with the long axis of the archoplasm. . . . 
These two peculiar forms of nucleoli are always to be found 
after the transition from the first into the second sperma¬ 
togenetic period, and throughout all the generations of the 
latter.” 

Pfliicke (’ 95 ), ganglion cells of Invertebrate/,: “ Ob . . . die 

zum Nucleolus tretenden Lininfaserchen mit der Substanz 
desselben verschmelzen oder jener dem Vereinigungspunkt der 
Geriistbalkchen nur aufgelagert ist, muss ich unentschieden 
lassen. Die Nucleolen erhalten sich hierin komplicirter als 
die Chromatinkornchen, und die Moglichkeit, dass der intensiv 
farbbaren Substanz des Kernkorperchens ein eigenes stiitzendes 
Liningeriist zu Grunde liegt, ist nicht ausgeschlossen.” Nucleo¬ 
lar vacuoles are normal structures, and are especially abundant 
in the cells of gasteropods ; he followed in life the process of 
the detachment of smaller vacuoles from a larger one, as well 
as the process of fusion of two vacuoles. In Helix “ kommen 
neben drei bis fiinf grosseren Hauptnucleolen mit einem oder 
mehreren Hohlraumen im Inneren, sehr zahlreiche ganz zer- 
streut liegende kleinere Nebennucleolen bis zur GrosSe eines 
Chromatinkornes herab vor, denen Vacuolen ganz fehlen und 
die sich von Chromatinkornchen nur durch die Farbung unter- 
scheiden.” He also observed (cells of gasteropods) the 
“ Kernkorperchenkreis ” first described by Eimer, and found 
that the circle of granules around the nucleolus was connected 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


with it by linin fibers ; but he was unable to decide whether 
these granules are thickenings of linin fibers, or whether they 
correspond to “Nucleolen bezw. Nebennucleolen . . . , welche 
sich vielleicht vom Mutterkorper getrennt haben und durch 
Wirkung centraler Lebensherde in jener typischen, regel- 
massigen Stellung verharren.” 

vom Rath (’ 95 a) studied the maturation of the ovum of 
Euchaeta marina: on PI. VII he figures a number of various 
sized, all rather large nucleoli, in the germinal vesicle, at the 
stage when the chromosomes are longitudinally cleft. 

vom Rath (’ 95 b) finds that the secretion of the gland cells 
of the head in Anilocra stains exactly like the nucleoli, and 
concludes that both substances are probably chemically related. 
He briefly mentions (footnote, p. 5) having seen double nucleoli 
in liver cells of molluscs and Amphibia; these dumbbell¬ 
shaped nucleoli may be either regarded as states of fusion or 
of division. In liver cells of Astacus the nucleolus consists of 
“ zwei verschieden tingirten einander dicht anliegenden Kugeln 
einer dunklen und einer blassen.” There is no relation between 
centrosomes and nucleolar substance. 

Rhumbler (’95) studied the nucleolar relations of Cyphoderia. 
From three to nine “ Binnenkorper ” lie within the nucleus, the 
largest nuclei having the smallest number; so that accompanying 
the increase in size of the nucleus, a gradual fusion of the 
“ Binnenkorper ” takes place, though without an appreciable 
increase in the total volume of their substance. 

Sacharoff (’95) concludes that since the eosinophilic granules 
of the blood have the same appearance as the nucleoli, “ und 
weil diese Kernkorperchen bei dem Herausfalien der Kerne auch 
herausfallen miissen, um dann unweigerlich von Leukocyten 
verschlungen zu werden, so ist mit grosster Wahrscheinlichkeit 
anzunehmen, dass bei Saugern die eosinophilen Granulationen 
auf dem Wege der Phagocytose von aus Hamatoblasten heraus- 
gefallenen Kernkorperchen entstehen.” In birds the nuclei 
do not fall out of the erythrocytes, but the eosinophilic cor¬ 
puscles are nucleoli which have wandered out of the nucleus; 
these nucleoli are rod shaped. (Only medical literature is cited 
in this paper.) 


MONTGOMERY. 


[Vol. XV. 


360 

Sala (’95), ovum of Ascaris: in the first maturation mitosis 
the single nucleolus breaks into small pieces of various size, 
which gradually become scattered throughout the nucleus ; 
then they become smaller and spherical, and come to lie directly 
under the nuclear membrane. These fragments may possibly 
stand in a genetic connection with the corpuscles which are 
subsequently found at each pole of the spindle. And since the 
latter corpuscles may stand in some connection to a centrosome, 
“ es ist . . . nicht unmoglich, dass eine enge Beziehung besteht 
zwischen der Auflosung des Nucleolus und dem Auftreten des 
Centrosoma.” 

Schloter (’95), gland and liver cells of Salamandra: in 
the nuclei may be distinguished, besides the chromatin and 
paralinin, red-staining spherical corpuscles, the larger of which 
are regarded as plasmosomes. 

Sobotta (’95), ovum of Mus: in contradiction to the view of 
Holl, the chromosomes are not derived from the nucleoli only, 
but from the whole chromatic substance of the nucleus. 

van der Stricht (’95) observed in the larger ovarial eggs of 
Amphioxus that each contains a large nucleolus with an excen- 
tric vacuole ; it disappears at the time of formation of the pole 
spindle. 

Vejdovsky (’95a) found large, homogeneous nucleoli in the 
yolk cells of Prorhynchus hygrophilus , “ die nicht die gewohn- 
liche kugelige Gestalt bewahren, sondern immer in Theilung 
begriffen sind. Man findet meist doppelte Kernkorperchen, 
deren Halften durch eine ziemlich tiefe Furche von einander 
getrennt sind und die eine centrale Hohlung erkennen lassen. 
Nebstdem findet man in Drei- selbst Viertheilung begriffene 
Kernkorperchen. . . . Ich glaube . . . , dass man es hier mit 
einer Hypertrophie der normalen Kernkorperchen zu thun hat, 
welche schliesslich zur Degeneration der Kerne fuhrt”; these 
nucleoli occupy more than two-thirds of the space within the 
nucleus. In the ovum the nucleolus is much smaller, and 
shows a division into two parts (Fig. 89), but here these two 
parts are not of equal size. 

Vejdovsky (’95b) found in the egg of Bothrioplana a spherical 
nucleolus, “mit einem Nucleolinus.” 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 361 

Waldeyer (’95), cited by Flemming (’96), regards the nucleoli 
as morphologically distinct from the chromatin reticulum. 

Wheeler (’95) observed in Myzostoma glabrum that the 
nucleolus is large and vacuolated, and after the reduction 
mitosis, “ remains in the cytoplasm as an inert mass, gradually 
melting away, but not disappearing until about the eight-cell 
stage, when it may often be found in the largest blasto- 
mere.” 

Wilcox (’95) holds that in the spermatocytes of Cicada the 
nucleoli stand in genetic connection with the centrosomes, 
and adds, “ It is probable that different structures have been 
called nucleoli by different authors.” 

i 8 g 6 . 

Auerbach (’96) studied the spermatogenesis of Paludina: 
the nucleus of the spermatogonium contains a number of 
large, more or less spherical bodies (“ Karyosomen ”); each 
nucleolus (of the resting spermatogonium), after simultaneous 
staining with acid fuchsine and methylen green, shows a 
central red portion and a blue peripheral shell. “ Es besteht 
also eine Zeit lang der Nucleolus aus einer erythrophilen Cen- 
tralmasse und einer kyanophilen Rinde.” In the subsequent 
nuclear division of these cells the nucleoli disappear. “ Fest 
steht nur, dass in dem Netzstadium die Nucleoli als solche 
verschwinden, und dass ihre Rindensubstanz auf die angegebene 
Art zu einem Teile des intranuklearen Netzwerkes wird, der 
anfangs noch unterscheidbar ist, dann aber durch Auseinander- 
riicken der Knotenpunkte sich in dem ubrigen Fadennetze ver- 
liert.” In the spirem stage there are one or two small, spherical, 
red-staining bodies in the nucleus ; he was unable to determine 
whether these stand in any genetic relation to the nucleoli, 
which had previously vanished. In the spermioblast (which 
changes directly into the hair-shaped spermatozoon) a small, 
red-staining body lies within the nucleus, but subsequently 
disappears ; Auerbach supposes that it wanders out of the 
nucleus and fuses with the “ Nebenkern.” 

Doflein (’96), maturation of the egg of Tubularia larynx: 
the single large nucleolus is suspended by achromatic fibers in 


3 62 


MONTGOMERY. 


[Vol. XV. 


a clear, structureless space within the nucleus ; at first homo¬ 
geneous, it later contains from one to five unstaining “ Korper- 
chen,” which he thinks are not vacuoles, on account of their 
refractibility. In the amitotic division of those nuclei which 
degenerate and eventually become absorbed by a definitive egg 
cell, division of the nucleolus precedes that of the nucleus. 

Floderus (’ 96 ) studied the maturation and embryonal develop¬ 
ment of various Tunicata. A “ Hauptnucleolus ” and “ Neben¬ 
nucleoli” are present. The former is homogeneous in only 
very young cells, and later differentiates into two different sub¬ 
stances : (i) a refractive, larger portion, which encloses (2) a 
less-refractive, paler portion. He considers the small vacuoles 
of the nucleoli to be “ Kunstprodukte,” though the large one is 
normal. “Nicht selten findet man in dieser grossen, allem 
Anscheine nach mit Fliissigkeit erfiillten Hohlung eine Anzahl 
fester, lichtbrechender Kornchen, vielleicht Coagulationspro- 
dukte, die wahrscheinlich bei der Fixierung entstanden sind.” 
As a rule there is one, but sometimes two “ Nebennucleoli ” 
in most though not all eggs ; these rarely attain half the 
diameter of the “ Hauptnucleolus,” and appear in the germinal 
vesicle shortly before the yolk granules arise in the cytoplasm; 
they are similar to, but paler than, the refractive portion 
of the large nucleolus. The “Nebennucleoli” are absent in 
Clavelina ; they probably arise by gemmation from the “ Haupt¬ 
nucleolus,” and he figures to this effect a lobular “ Hauptnu¬ 
cleolus.” In the cytoplasm of the ova of Styelopsis and Ciona 
(but not Clavelina and Corella) certain spherical “ intravitelline 
Korper” occur, usually one to a cell, and frequently close to 
the nuclear membrane ; in size and staining reactions these are 
similar to the “ Nebennucleoli,” and, following Roule, “ sehe ich 
mich genothigt, anzunehmen, dass sie von Nebennucleolen 
herriihren, die aus dem Kern des Eies in den Dotter hinausge- 
wandert sind,” thereby supposing that they press out through 
a preliminarily produced pore in the nuclear membrane, and 
that the larger intravitelline bodies are probably fused masses 
of smaller ones. In accord with Henneguy (’ 93 ) and Roule he 
considers the intravitelline bodies not as “ Dotterkerne ” nor 
astrospheres, but as atavistic or rudimentary organs, which 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 363 


together with the nucleoli correspond to the macronucleus of 
the Infusoria. 

Gerould (’96), ovarial eggs of Caudina: in the youngest ova 
there are numerous peripheral nucleoli; these increase in size 
as the nucleus grows, and subsequently each contains a vacuole, 
but they are always close to the nuclear membrane. 

Greenwood (’96), macronucleus of Carchesium polypinum: 
the nucleoli (“ protomacrosomes,” in distinction to the “pro- 
tomicrosomes,” or chromatin granules) are numerous and 
vacuolated, and stain like true metazoan nucleoli. They vary 
in size and form, and are probably amoeboid, though this point 
could not be determined in the living nucleus, which is first 
rendered visible by reagents. The vacuoles are fluid accumula¬ 
tions, and arise first in the center of the nucleolus. “No 
vacuoles surround the macrosomes of Carchesium at any time, 
nor do they ever show general increase of fluidity or swelling 
such as might accompany the penetration through them of some 
secretion from without ; . . . the deposition of vacuolar fluid is 
centrifugal ; . . . thus the macrosome may become a bladder¬ 
like or honey-combed structure, its residual solid (?) forming a 
well-defined membrane-like investment for fluid contents.” 

Henneguy (’96) distinguishes true and false nucleoli (the 
latter being “ noeuds du reseau,” in the sense of O. Hertwig, 
’92). He reviews the observations upon nucleoli made by 
several previous authors. 

R. Hertwig (’96), unfecundated ova of echinoderms poisoned 
with strychnine : the nucleoli vanish within the nucleus as the 
chromosomes appear. “ Meine eigenen Untersuchungen lassen 
es mir ausgeschlossen erscheinen, dass im Ei der Seeigel Nu- 
cleolen und Centrosomen irgend etwas mit einander zu thun 
haben. . . . Dagegen ergeben sich unzweifelhafte Beziehungen 
der Nucleoli zur Entwicklung der Chromosomen. . . . Dieses 
Wechselverhaltniss ist nun nicht so zu verstehen, als ware das 
gesammte Material der Chromosomen in den Nucleoli enthalten. 
Dagegen spricht die geringe Masse der Nucleolar-Substanz und 
ihr verschiedenes Verhalten den iiblichen Chromatin-Farbungs- 
mitteln gegeniiber. . . . Die Nucleolen konnen somit den 
Chromosomen ein zur endgiiltigen Fertigstellung nothwendiges 


3 6 4 


MONTGOMERY. 


[Vol. XV. 


Erganzungsmaterial liefern.” “Chromatin-Nucleoli” are such 
as contain the whole chromatin of the nucleus ( Actinosphae - 
rium, Spirogyra, salivary glands of Culex ); “ solche Kerne 
zeigen dann ein achromatisches Gerust und in demselben einen 
grossen chromatischen Korper, im iibrigen Nichts, was man den 
Nucleoli oder den Chromosomen der Gewebszellen vergleichen 
konnte. . . . Derartige Nucleoli waren dann nicht, wie mein 
Bruder annimmt, und auch ich friiher geglaubt habe, von 
den echten Nucleoli als etwas wesentlich Verschiedenes zu 
unterscheiden ; sie wiirden Nucleoli sein, die ausser der specifi- 
schen Nucleolensubstanz noch das Chromatin des Kernes 
enthalten. . . . Bei der Umwandlung zur Spindel losen sich 
Chromatinkornchen vom Nucleolus ab und treten auf das 
Kernnetz iiber, ein Substrat hinterlassend, das man wohl den 
echten Nucleolen vergleichen muss. Spater wird auch dieses 
aufgelost ” 

Korschelt (’ 96 ), employing a modification of the Ehrlich- 
Biondi stain, finds in the spinning glands of caterpillars that the 
macrosomes stain green and hence consist of chromatin, while 
the microsomes stain red and so must be regarded as nucleoli 
(cf. ’97). 

List (’ 96 ) made comparative studies on various nucleoli, 
principally with a view to their chemical constituents, by apply¬ 
ing a new staining method, whereby Berlin blue is produced in 
the fixed tissues. “Wir sind zu dem Resultate gekommen, 
dass die Nucleolarsubstanzen nach ihrem chemischen Verhalten 
3 verschiedene Gebilde darstellen, von denen jedes wahrschein- 
lich wieder eine eigene complicirte chemische Zusammensetzung 
besitzt. Nach der bisherigen Bezeichnungsweise sind zu unter¬ 
scheiden : Hauptnucleolus, Nebennucleolus und Nucleolus 
schlechtweg the substance of all the nucleoli differs from 
that of the nuclein (chromatin) proper. “ Wir haben gesehen, 
dass (bei Mytilus und Pristiurus) die Umsetzung des Ferro- 
cyankaliums durch Salzsaure, wodurch Ferrocyanwasserstoff- 
saure und hieraus durch den Sauerstoff der Luft Berlinerblau 
entstand, allein geniigte, um die Nebennucleolen zu farben. . . . 
Wenn wir die Reagentien concentrirter anwenden, ... so tritt 
in jeder Zelle die Substanz des Nucleolus in Gestalt eines oder 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 365 


mehrerer blauer Kiigelchen hervor. . . . Nach ihrem chemischen 
Verhalten stehen also Nebennucleolus und Nucleolus einander 
naher als Haupt- und Nebennucleolen ”; he concludes that the 
“Nucleolus” and the “Nebennucleolus . . . mindestens ver- 
schiedene Modificationsstufen des Paranucleins . . . darstellen.” 
Mytilus egg: what Lonnberg supposed to be vacuoles within 
the nucleoli, List holds are “ Nebennucleoli,” and these alone 
evince the characteristic Berlin-blue reaction ; by afterwards 
staining the preparation with carmine, “ die Masse des Haupt- 
nucleolus, das Nuclein, hatte sich scharf roth gefarbt, die 
Nebennucleolarsubstanz, das Paranuclein, rein blau.” Even in 
eggs where no yolk was as yet present, both these substances 
could be demonstrated. The “ Hauptnucleolus ” represents 
the greater part of the nucleolus, and is usually single ; in it 
may lie one spherical “ Nebennucleolus,” or the latter may 
cover, cap-like, one pole of the former; sometimes “Nebennucle¬ 
oli ” occur in the nuclear cavity, separated from the “ Haupt¬ 
nucleolus ”; occasionally there are true vacuoles within the 
latter. Pholas egg: (treatment with iron chloride, nitric 
acid, then “ Ferrocyankaliumlosung”) the “Nebennucleolus” 
is much larger than the “ Hauptnucleolus,” except in very 
small ova, where they may be equal in size. The last-named 
nucleolus may enclose a large, excentric vacuole, or in place 
of this, a “Nebennucleolus”; in the chromatin network of 
the nucleus there are small nodules of paranuclein, and some¬ 
times a free “Nebennucleolus.” “In alteren Eiern iiberwiegt 
bei Weitem der Nebennucleolus den Hauptnucleolus an Masse ”; 
the latter is either apposed to one end of the former, or there 
may be a large “Nebennucleolus” with a small “ Hauptnucle¬ 
olus ” at each end of it. Pristiurus egg : in the youngest 
germinal vesicles the minute nucleoli all lie at the nuclear 
periphery, the larger ones being central; in larger ova all the 
nucleoli are placed at the periphery of the nucleus. Sphaere- 
chinus egg : the supposed (Hacker, ’ 93 b) vacuole of the “Neben¬ 
nucleolus ” is in reality the “ Hauptnucleolus ”: “ Jedoch weichen 
meine Resultate von denen Hacker’s darin principiell ab, dass 
eben festgestellt werden konnte, dass das, was H. Hauptnucle¬ 
olus nennt, wie ein Nebennucleolus reagirt, und die Vacuole 


366 


MONTGOMERY. 


[Vol. XV. 


wie ein Hauptnucleolus.” With the three staining methods 
employed (all used on material fixed with corrosive sublimate), 
only the “ Nebennucleolus ” is plainly stained, “nicht aber der 
Nucleolus schlechthin, wie er in jeder Zelle vorkommt.” By 
treatment for half an hour with a drop of .5 Jo iron chloride 
solution, then stained by the Berlin-blue reaction, in each 
somatic cell the nucleolar substance appears in the form of 
bluish-green spherules. “Im Mollusken- wie im Vertebraten- 
gewebe hatte jede Zelle einen rundlichen Nucleolus; in secer- 
nirenden Zellen, z. B. Darmzellen, traten 2 oder 3 auf, oder 
Grossenunterschiede, wie z. B. der Nucleolus in der Leberzelle 
von Mytilus durch seine Grosse auffallt.” 

Michel (’ 96 ), ova of Nephthys and Spiophanes: each double 
nucleolus consists of (1) a darker, more granular, portion, which 
in Spiophanes contains either a small granule or a vacuole (he 
is undecided which it is), and in Nephthys is vacuolated; and 
(2) of a clearer, refractive, unstaining portion. In Nephthys 
there are usually two double nucleoli, “ la substance colorable 
recouvrant plus ou moins completement la masse claire comme 
d’une calotte but other states were also found : “trois nucle- 
oles doubles, une sphere claire entre deux parties sombres 
presque a l’oppose; inversement, une partie sombre et deux 
spheres claires presque opposees, nucleoles plus composes avec 
plusieurs spheres claires et meme comme spumeux, spherules 
claires libres en plus de celles des nucleoles doubles jusqu’a 
une douzaine. . . . Les masses claires, avec leur aspect, leur 
forme spherique et leur deformation temporaire par la pression, 
leur variation de taille suivant les conditions osmotiques, 
l’epaississement de leur paroi par reduction de volume, apparais- 
sent comme des vesicules a contenu liquide special,” while the 
colorable portions are composed of pyrenin, and hence are true 
nucleoli (the pyrenin proved “ par l’absence de gonflement par 
1’eau et par le gonflement par les acides, par l’insolubilite dans 
le sulfate de cuivre ou le ferrocyanure de potassium. . . . 
l’aspect des vesicules et leur disposition dans les nucleoles ou a 
Y 6 tat libre . . . portent a croire a des vacuoles a contenu special 
formees dans le nucleole et finalement eliminees ”). 

Morgan (’ 96 ) studied Echinoderm eggs placed in artificial 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 367 

media: immature ova of Sphaerechinus , placed in sea water to 
which 1.5 gr. NaCl had been added, show artefacts in the 
nucleolus: “ Each [body] consists of an outer darker shell, 
which is filled with a clear fluid, and the center of each sphere 
is occupied by a small black granule”; several of these struc¬ 
tures are usually found on each section through the nucleolus. 
(For previous descriptions of somewhat similar productions, cf. 
Ransom (’ 67 ), Leydig (’88), and O. Schultze (’ 87 ). The upper 
of the two figures numbered “24” in Morgan’s plate should 
be “23,” since it refers to the nucleolus.) 

Rohde (’ 96 ), ganglion cells of Doris and Pleurobranchus : the 
nucleoli wander out of the nucleus and finally into the neuroglia, 
and there acquiring an envelope (derived from the neuroglia) 
form new cells. [Judging from his figures, however, these 
supposed nucleoli would seem to be myelin drops.] 

Wagner (’ 96 a), spermatogenesis of Arachnids: “ Bei der 
ersten Spermatocytentheilung theilt sich der Nucleolus ent- 
weder in der Ebene der Aequatorialplatte mit den Chromo- 
somen zusammen, oder ausserhalb derselben neben einem der 
Spindelpole. Im letzteren Falle tritt er nach dem Ver- 
schwinden der Kernhiille . . . aus dem Kerne heraus.” 

Wheeler (’ 96 ) gives no description of the nucleoli in the text, 
but he figures several stages of the development in eggs of 
Myzostoma (Figs. 9, 10-1$, M. cirriferum ; Figs. 23, 52-54, 56, 
M. glabruni). In M. cirriferum (Figs. 12-15) * s figured, in 
addition to the single large nucleolus, also one smaller nucleolus. 

E. B. Wilson (’ 96 ) states of the true nucleoli or plasmo- 
somes : “ There is strong evidence that the true nucleoli are 
relatively passive bodies that represent accumulations of reserve- 
substance or by-products, and play no direct part in the nuclear 
activity.” In germinal vesicles he assumes that the “ principal 
nucleolus ” is chemically different from the nucleoli of somatic 
cells ; but that the “ accessory nucleoli ” of the former corre¬ 
spond to the nucleoli of the latter. He concludes that “ we can 
hardly doubt the conclusion of Hacker, that the nucleoli of the 
germ-cells are accumulations of by-products of the nuclear action, 
derived from the chromatin either by direct transformation of 
its substance, or as chemical cleavage-products or secretions.” 


368 


MONTGOMERY. 


[Vol. XV. 


1897. 

Toyama, cited by R. Hertwig (’ 96 ), holds that the nucleoli 
become centrosomes in the spermatogenesis of Bombyx. 

Van Bambeke (’97a), ovocyte of Pholcus: there is usually a 
single large nucleolus, rarely also accessory ones ; the nucleolus 
is vacuolated, “ les vacuoles . . . faisant frequemment saillie a 
sa [tache germinative] surface; dans certains vacuoles, on 
decouvre des granules safraninophiles.” At a later stage the 
nucleolus retains much the same appearance, “ mais frequem¬ 
ment le contour net, safraninophile, qui la delimitait, a disparu 
en tout ou en partie, et l’on remarque parfois une solution 
decontinuite au niveau de laquelle le contenu de la tache 
s’epanche dans le reste du contenu nucleaire. Cette sorte 
d’evacuation ne doit pas etre confondue avec la rupture de 
vacuoles nucleolaires, laquelle peut s’observer a tous les stades.” 
(’ 97 b, the same, with figures.) 

Bouin (’ 97 ), giant spermatogonia of Cavia: the accessory 
part (“corps juxtanucleolaire ”) of the double nucleolus stains 
red in safranine and blue in haematoxylin (in opposition to 
Hermann), though less deeply than the spherical portion of the 
nucleolus, and is sometimes hemispherical in form. This part 
is single, and appears to consist of a mass of very fine granules. 
In degenerating cells, “ les uns nous montrent deux nucleoles 
flanques chacun d’une ou de plusieurs petites masses hemi- 
sphdriques, refringentes et teintees en rose pale ; lors des 
mouvements intranucleaires, les corps juxtanucleolaires con- 
tractent des rapports plus intimes avec les vrais nucleoles, 
deviennent plus refringents et moins colorables, s’accolent a 
leur substance, se divisent a leur suite, et les accompagnent 
dans leurs migrations. Apres plusieurs divisions repet^es, ces 
noyaux contiennent un certain nombre de nucldoles, cinq ou six 
generalement.” In the process of formation of the cells of 
Sertolli the nucleoli fuse successively with one another. 

Braem (’ 97 ), Plumatella: in the egg of .013 mm. diameter 
the nucleolus contains one to four vacuoles: “ Sie sind allem 
Anschein nach Fliissigkeitsblaschen, welche im Nucleolus auf- 
treten und auf dem Hohepunkt ihrer Entwickelung an die 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 369 


Peripherie riicken, um da ihren Inhalt nach aussen zu ent- 
leeren.” The nucleolus becomes ovoid, and its substance 
paler at its smaller end; the vacuoles are usually, but not 
always, at the paler end. “ Zuweilen ist der Gegensatz der 
beiden Nucleolus-Halften lediglich in der verschiedenen Farb- 
barkeit derselben ausgesprochen. In anderen Fallen wird er 
durch eine Einschniirung bezeichnet, die den Nucleolus in 
einen grosseren, dunkeln und einen kleineren, hellen Abschnitt 
zerlegt. . . . Die Einschniirung kann nun zu einer volligen 
Abschniirung fiihren, so dass^ der Nucleolus doppelt erscheint 
und von zwei neben einander liegenden Kugeln gebildet wird, 
oder bei gegenseitiger Entfernung der Theilstiicke in zwei 
raumlich getrennte Nucleoli zerfallt. . . . Selten ist der Nucle¬ 
olus dreitheilig .. ., wo das mittelste Stuck dunkler ist als die 
beiden seitlichen... . Dies lasst vermuthen, dass der Keimfleck 
im Stande ist, unabhangig vom Wachsthum des Eies seine 
Gestalt zu verandern, und dass die Zweitheiligkeit auf der 
Bildung eines pseudopodienartigen Fortsatzes beruht, der sich 
bald mehr, bald weniger deutlich vom Hauptkorper abgegliedert 
und auch hinsichtlich seiner Substanz bald mehr, bald weniger 
von demselben verschieden ist.” 

De Bruyne (’97), double cells of the ovarian follicle of Nepa , 
Periplanetciy Meconema , and Aeschna: in the amitotic division 
of the nucleus the nucleolus divides first. (Since the cytoplasm 
does not divide, each such cell finally receives two nuclei.) 

Carnoy and Lebrun (’97a) (an abstract of this paper may also 
be found in the American Naturalist for July, 1897). This con¬ 
tribution deals particularly with the relations of the nucleoli in 
the growth period of the ovum of Salamandra and Pleurodeles. 
In the youngest nuclei observed there is a nuclein filament, 
but no nucleoli; the first nucleoli arise as buds from the fila¬ 
ment, and these are termed “ nucleoles primitifs.” Then the 
nuclear filament becomes changed into an amorphous magma, 
composed of irregular granules, and the latter then subse¬ 
quently disappear, so that all trace of the original filament 
becomes lost. All further changes within the nucleus are 
of nucleolar character. From the “nucleoles primitifs” are 
derived the “ nucleoles secondaires ” which “ sont dus a des 


37° 


MONTGOMERY. 


[Vol. XV. 


associations de granules provenant de la desagregation de 
I’Element nucldinien ” ; and then follow the “ nucleoles ter- 
tiaires,” which differ from the nucleoli of the preceding two 
generations in that they do not come from degenerating gran¬ 
ules of preceding generations, but are detached from them in 
the form of spherules. Each nucleolus of each generation 
arises, increases in size, becomes more complex in structure, 
and then passes through a polymorphic “ figure de resolution ” ; 
the form of these figures varies according to the particular 
generation, and also according to particular ova. The greater 
part of the “ figure de resolution ” then disappears, except a few 
granules which serve as the starting point for the next genera¬ 
tion ; that portion of the substance which disappears serves as 
nourishment for the egg. So all the generations of the second¬ 
ary and tertiary nucleoli arise “a l’aide des produits de la 
resolution anterieure.” After each “ resolution ” new nucleoli 
arise, and the number of these generations is large, continuing 
through a length of three years. The number of primary 
nucleoli is usually from two to six ; of secondary, from 400 to 
500 ; of tertiary, from 500 to 1000 ; the number varying in 
different ova. Fusions of nucleoli are of normal occurrence : 
“cette attraction des masses nucleiniens rappelle a 1’esprit ce 
qui se passe au sein de Toeuf entre les noyaux de conjugaison.” 
In the radiation exerted by each nucleolus upon the surrounding 
caryoplasm “nous voyons ... la confirmation d’une th&se 
soutenue dans la ‘ Cytodierese/ a savoir : que c’est sous l’influ- 
ence du noyau que se forment les asters de division.” The 
chromatin filament does not reappear, but there is a “grand 
nombre de generations nucleolaires et filamenteuses qui naitront 
et disparaitront tour a tour, Tune apres l’autre, jusqu’a l’epoque 
des globules polaires.” The authors necessarily regard all the 
previous observations on the amphibian ovum as erroneous. 
General conclusions for all kinds of cells, based in part on 
previous observations : there may be distinguished “nucldoles 
plasmatiques,” “nucleoles nucldiniens,” and “nucleoles mixtes ” 
(“qui sont rare”). Plasmatic nucleoli consist of at least two 
substances, “ une plastine et une globuline digestible.” All 
nucleoli, “ lorsque leur formation est achevee . . . represented 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 371 

la totalite de l’dlement filamenteux d’un noyau ordinaire ; . . . 
dans bien de cas — aujourd’hui nous pourrions peut-etre dire 
dans tous — on constate dans ces corps la presence d’un veri¬ 
table appareil filamenteux, tortille sur lui-meme, comme dans 
un noyau ordinaire, et presentant les memes proprietes que 
dans ces derniers. C’est que Ton voit surtout dans les nucle- 
oles-noyaux, c’est-a-dire dans les nucleoles nucleiniens uniques, 
qui ont absorbe tout l’el^ment filamenteux primitif.” All 
nucleoli develop from the chromatin filament ; and chromo¬ 
somes are derived from “ nucldoles noyaux.” The chemistry 
of nucleoli is also considered. 

Carnoy and Lebrun (’ 97 b), fecundation of the ovum of 
Ascaris megalocephala: the centrosomes are “ nucleoles plas- 
matiques ou achromatiques ” which have left the nucleus at the 
commencement of mitosis ; one is derived from the male, the 
other from the female, nucleus. They totally disappear after 
mitosis, and neither reenter the nuclei nor divide to produce the 
centrosomes of the subsequent division. 

Cunningham (’97) : “ There are indications in the ova of the 
turbot that the substance of the nucleoli is absorbed into 
the central fibrils to form the chromosomes of the polar mito¬ 
ses, but the actual formation of these chromosomes was not 
followed.” 

v. Erlanger (’97), a brief mention of certain recent views 
upon the nucleolus : “Als echte Nucleolen waren allein solche 
Korper zu bezeichnen, welche sich durch ihr Verhalten gegen 
Chemikalien ... scharf von dem Chromatin unterscheiden. 
. . . Vorderhand bleibt also die Bedeutung der echten Nucle¬ 
olen ratselhaft, falls man diese Gebilde nicht mit Hacker als 
eine Sekretion des Kernes beurteilen will.” They bear no 
relation to centrosomes. 

Fauvel (’97), ovogenesis of Ampharete: ovarial ova of 30/u 
diameter, and at this stage only, contain two nucleoli. “ On 
rencontre toutes les modifications : nucleole simple, nucleole 
etrangle par le milieu, deux nucleoles accoles, et enfin deux 
nucleoles bien nettement separes. . . . Nous n’en avons 
jamais rencontrd deux dans l’oeuf mfir, ni dans l’oeuf non 
d&ache de l’ovaire.” The nearly mature ovum contains one 


372 


MONTGOMERY. 


[Vol. XV. 


large nucleolus, with a large vacuole ; he believes that subse¬ 
quent to the two-nucleolus stage one of the nucleoli is extruded 
from the nucleus. Two nucleoli were observed also in the ova 
of Amphicteis , Sanytha , and Melinna. 

Flemming (’97) recurs to the controversy between Korschelt 
(’ 96 ) and Meves (’97), and agrees with Meves that the macro- 
somes are nucleoli, and the microsomes chromatin granules. 
He also mentions the following observation on the ovum of 
Ascidia ca?iina: here there is one “ Nucleolus ” and one much 
smaller “ Kernkorper ” ; “ beobachtet man ihn [Kernkorper] 
am lebend entnommenen Ei, so findet man ihn so gut wie 
stets in Molekularbewegung, und zwar oft in recht grossen 
Exkursionen.” 

Hacker (’97a) (’96 is a preliminary communication), cleavage 
stages of Cyclops brevicornis. This paper deals particularly with 
the “ intraspharale,” “ extranucleare,” or “ Aussen-Kornchen 
(Ektosomen) ” found in certain of the astrospheres of the 
cleaving ovum. These ectosomes are small spherical bodies of 
various size, which stain like the nucleoli, but somewhat more 
intensely. In the resting stage of the cell there are several 
nucleoli in the nucleus, and no ectosomes outside of it ; when 
the nucleus enters on the aster stage, the nucleoli have disap¬ 
peared and ectosomes are present in one of the astrospheres, 
at first at the base of, subsequently on the whole periphery of, 
the latter ; towards the close of the metakinesis there appear 
in the place of. the ectosomes larger clumps of red-staining 
substance. He concludes : “ So glaube ich es denn mit Sicher- 
heit aussprechen zu diirfen, dass diese groberen Brocken auch 
genetisch mit den Kornchen [Ektosomen] zusammenhangen, 
sei es, dass sie direkte Umwandlungsprodukte derselben, sei es, 
dass sie Neubildungen sind, welche dem namlichen Process 
ihre Entstehung verdanken, aber in Folge der wahrend der 
Theilung eintretenden Zustandsanderungen der Zelle eine etwas 
andere Beschaffenheit, einen anderen Aggregatzustand ange- 
nommen haben. Wie ich gleich hier hinzufiigen mochte, ver- 
schwindet die Erscheinung, sowohl im Zweizellenstadium als in 
den spateren Stadien, wahrend der eigentlichen Ruhepause 
vollstandig, indem vermuthlich jene Massen einer Resorbtion 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


n *7 'j 
0/0 

Oder chemischen Umwandlung anheimfallen.” In only one 
astrosphere of only one cell in each of the following cleavage 
generations this process is repeated, and the line of these par¬ 
ticular cells (“ Kornchen-Zellen ”) constitutes the line of devel¬ 
opment of the sexual cells ; but the ectosomes are present in 
these particular cells only during mitosis, and in the resting 
stages are absent, while nucleoli occur in the nuclei ; this proc¬ 
ess was observed from the first through the ninth cleavage 
stages. He concludes that in each generation there is a produc¬ 
tion de novo and a subsequent solution (“Auflosung ”) of the 
ectosomes. The first appearance of the latter coincides in point 
of time approximately with the disappearance of the nucleolar 
substance in the nucleus ; from this and certain other factors 
he concludes : “ So . . . wiirde also der Annahme kaum etwas 
im Wege stehen, dass die zu Beginn der Mitose noch vorhan- 
denen oder neugebildeten Nucleolen aus dem Kernraum in der 
Richtung der einen Spahre auswandern und sich hier in die 
Aussenkornchen umwandeln. ... Fur die Kerne der Korn- 
chenzellen ist dann allerdings, in Gegensatz zu den ubrigen 
Embryonal-Elementen, eine besonders reichliche Produktion 
der Nucleolarsubstanz und demnach eine besonders intensive 
vegetative Thatigkeit [cf. ’95] anzunehmen.” The explanation 
for the arrangement of the ectosomes in only one of the 
astrospheres he finds in the assumption “dass die beiden 
Centrosomen einen verschiedenen (vielleicht einen verschieden 
* kraftigen ’) Einfluss auf das umgebende Plasma, beziehungs- 
weise auf die beweglichen Inhaltskorper desselben ausiiben.” 

Hacker (’ 97 b) finds that in germ cells of both animals and 
plants there is to be noted “das Auftreten eines einzigen, 
vacuolenhaltigen, dunkel tingierbaren “ Hauptnucleolus ” in 
den jiingeren Stadien, das Hinzutreten von blasseren adventiven 
oder “ Neben-Nucleolen ” in einer friiheren oder spateren 
Phase.” Nucleolar substance arises during one or several stages 
of nuclear activity as a by-product of metabolism, possibly also 
as chromatin substance which has become structureless and 
chemically changed; and, finally when the nucleus begins to 
divide, is removed out of the latter. He confirms Wheeler’s 
(’ 96 ) observations on the ovum of ATyzostoma, that the nucleolus 


374 


MONTGOMERY. 


[Vol. XV. 


wanders out of the nucleus into the cytoplasm, where it slowly 
decreases in size. 

Hermann (’ 97 ) figures (Fig. 20 ) a spermatogonium nucleus of 
Scyllium containing a single and a double nucleolus ; the latter 
consists of two apposed spheres, which differ chemically and 
dimensionally. 

Korschelt (’ 97 ) maintains his previous opinion (’96) of the 
chromatin nature of the macrosomes of the nuclei in the 
spinning glands of caterpillars, in answer to the criticism of 
Meves (’ 97 ) (reviewed immediately below). Korschelt employed 
the Ehrlich-Biondi stain with increased strength of the methyl 
green, and thereby obtained a coloration of the macrosomes 
and microsomes the very opposite of that procured by Meves. 
“ Ob man iiberhaupt achromatische, chromatische Substanz und 
Nucleolen in alien Kernen so scharf auseinanderhalten kann, 
wie dies vielfach geschieht, ist mir hochst zweifelhaft. Wenn 
man in verschiedenen Zustanden der Kerne Nucleolen auf- 
treten und wieder verschwinden sieht, wird man annehmen 
miissen, dass sie sich aus den sogenannten achromatischen oder 
chromatischQn Substanzen des Kerns, vielleicht aus beiden 
herausbilden. So konnen sich moglicher Weise auch die von 
mir als Makrosomen bezeichneten Theile in Nucleolen umbilden 
und das von Meves angegebene Auftreten von Vacuolen in 
ihnen wiirde damit seine Erklarung finden.” 

Meves (’ 97 ) contends that the microsomes in the spinning 
glands of caterpillars, which Korschelt regarded (’96) as lanthanin 
granules, are chromatin; and what Korschelt regarded as chro¬ 
matin granules (i.e., the macrosomes) are nucleoli. Meves 
employed the usual formula of the Ehrlich-Biondi stain (Heiden- 
hain’s receipt), and finding that the macrosomes thereby become 
stained red, concludes from this reaction their chromatin nature. 

Stauffacher (’97) finds in the aster stage of the mitosis of one 
of the pronephral cells of Cyclas , that the nucleolus still persists 
intact in apposition to the spindle fibers. 

Wheeler (’97), maturation of the ovum of Myzostoma: this 
object, previously described by the author (’ 95 ), is here more 
fully described with the addition of figures. A remarkable 
mode of formation of nucleoli in the pronuclei is described ; 


) 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


375 


each “ chromosome ” consists of “ two granules, at first of the 
same size [which] grow very unequally, so that one is often 
considerably larger than the other. Hereupon some, but not 
all, of these granules break down to form irregular strings of 
minute karyomicrosomes which are distributed along the fibers 
of the achromatic reticulum. . . , The large chromatin granules 
which do not break down become the nucleoli of the pronuclei. 
I am unable to state positively that in each Diplococcus-shaped 
chromosome one of the granules breaks down to form a chain 
of minute karyosomes while the other persists as a nucleolus, 
but I am very strongly inclined to believe that this is the case.” 
These nucleoli are cast out into the cytoplasm when the first 
cleavage spindle is formed, and there rapidly dissolve. Wheeler 
accepts “Hacker’s view of the secretory nature of the nucleo¬ 
lus, at least so far as the germinal vesicle is concerned.” 

Bancroft (’ 98 ), germinal vesicle of Distaplia: the nucleolus 
“does not form the stellate body found in the old ova, as 
Davidoff maintained, but is found within this body, which is 
itself the remains of the germinal vesicle. The nucleolus at 
this stage is quite complex, consisting of a homogeneous cortex, 
an excentric finely granular medulla, and within the latter 
several very highly refractive bodies, the largest of which may 
have a granular appearance. During the greater part of the 
growing period these refractive bodies are the only substance 
in the germinal vesicle that takes the chromatin stain with a 
methyl green and acid fuchsine combination.” 

1898. 

Kostanecki (’ 98 ) confirms the observations of Wheeler (’ 95 , 
’97) in regard to the casting out of the nucleolus into the 
cytoplasm, in the maturation of the ovum of Myzostoma. 

B . Botanical Literature. 

Schleiden (’ 38 ) is the discoverer of the nucleolus in plants, but 
he gives it no name : “ einen kleinen, sich scharf abgren- 
zenden Korper, der, nach dem Schatten zu urtheilen, einen 


376 


MONTGOMERY. 


[Vol. XV. 


dicken Ring oder ein dickwandiges hohles Kiigelchen darzu- 
stellen scheint while in other cases it may be a simple spot, 
or may be wholly absent. “Aus meinen Beobachtungen an 
alien Pflanzen, die eine vollstandige Verfolgung des ganzen 
Bildungsprocesses erlaubten, geht hervor, dass dieser kleine 
Korper selbst friiher sich bildet, als der Cytoblast [Nucleus].” 

Macfarlane (’ 81 ) examined various plant cells, in all of which 
he found one or several bodies (“ nucleolo-nuclei ”) within the 
nucleolus. The nucleolus of Spirogyra has a distinct membrane, 
which disappears at the period of the nucleolar division ; the 
karyokinesis results in the formation of a “nuclear barrel,” at 
each end of which is a mass of nucleoplasm, these two masses 
being connected by fibers with the nucleolus which lies between 
them. The nucleolus then divides, preceded by a division of 
the nucleolo-nucleus, so that each daughter-nucleolus receives 
a daughter nucleolo-nucleus, and the daughter-nucleoli then 
wander apart to the nearest masses of nucleoplasm, “ as they 
retreat from each other they drive the polar masses before them, 
thereby elongating the nuclear barrel. . . . The nucleoli at 
length advance to the polar masses and bury themselves in 
the nucleoplasm of these.” From these and numerous other 
observations, Macfarlane concludes : “that the nucleolus, or 
more probably the nucleolo-nucleus, is the center of germinal 
activity, and that as we pass outwards to the periphery of the 
cell, this reproductive activity becomes less and less. In no 
other way, to my mind, can the number of nucleoli and nucleolo- 
nuclei at different ages in the cells of any plant be explained.” 

Strasburger (’ 82 a) gives reviews of previous observations on 
the chemical constituency of nucleoli. 

Strasburger (’ 82 b) studied nuclear division in various plant 
cells ( Fritillaria , Lilium, Hemerocallis , Tradescantia , Galanthus , 
Dicotyledons). “ Pollenmutterzelle ” of Fritillaria : between 
the nucleus and its membrane collects a homogeneous, refrac¬ 
tive, lens-shaped mass of substance; “ sie geht nicht unmittelbar 
aus den Kernkorperchen hervor, die ja schon auf vorausgehenden 
Stadien verschwunden waren, vielmehr reprasentirt sie, allem 
Anschein nach, ein Secret”; this body he terms “ Secretkor- 
perchen.” At first it stains deeply with methylen green ; but 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


377 


subsequently it ceases to stain, vacuoles arise in it, and it 
decreases in size, until at the time of the spindle formation it 
disappears. “ Sie [Secretkorperchen] treten erst auf, nachdem 
das Kernkorperchen oder die Kernkorperchen in dem Faden- 
knauel des Kerns Aufnahme gefunden. Ihre Entwicklungsge- 
schichte unterscheidet sich auch von derjenigen echter Nucle- 
olen, denn sie treten nicht im Verlauf der Fadenwindungen auf, 
vielmehr ausserhalb derselben, stets an der Wand der Zelle. 
Ausgeschlossen ist ja nicht, dass in der so ausgesonderten 
Substanz die Substanz friiher Kernkorperchen vertreten sei, 
aber erweisen lasst sich dies nicht.” So he concludes that 
before the mitosis of the spores and “ Pollenmutterzellen ” a 
certain change occurs in the nucleoplasma, in connection with 
the formation of the “ Secretkorperchen.” The nucleoli of 
many plant cells contain vacuoles. In the embryo sac of 
Galanthus a division of the large nucleolus takes place, which 
division is probably passive, caused merely by the tension of the 
cytoplasm. Gradations are to be found between the nucleoplas- 
mic-microsomic substance and the substance of the nucleoli : 
“ob die Nucleolen-Substanz trotzdem nur eine Modification 
der Microsomen-Substanz sei und aus dieser hervorgehe, will 
ich dahingestellt bleiben lassen. Wahrscheinlich ist mir aber 
das letztere, wenn ich bedenke, dass bei Eintritt in die Thei- 
lungsvorgange selbst die stark modificirte Nucleolen-Substanz 
in das Kerngerust findet und sich in demselben nicht anders 
als wie die Mikrosomen-Substanz verhalt. Man konnte die 
Nucleolen-Substanz vielleicht als einen Reservestoff des Zell- 
kerns auffassen, als eine momentan ausser Aktion gesetzte 
Substanz.” 

Tangl (’ 82 ) studied the nuclear division of three species of 
plants. Hemerocallis fulva , flower buds : the “ Pollenmutter- 
zelle” contains at first three or five nucleoli, which are homo¬ 
geneous. “ Mit fortschreitender Entwicklung der Mutterzellen 
verringert sich die Anzahl der Nucleolen,” until only one is to be 
found; this one is always peripheral in position, never in contact 
with the central “ Kornermasse.” Later, vacuoles appear in the 
nucleolus (he believes these to be the results of reagents), and 
while it still stains with carmine it no longer does with acidified 


378 


MONTGOMERY. 


[Vol. XV. 


methylen green. In mitosis, when the nucleus is uninucleolar, 
the substance of this nucleolus becomes dissolved in the nucleus; 
when multinucleolar, however, one of the nucleoli may pass 
out into the cytoplasm. Hesperus, “ Pollenmutterzelle ”: here 
there is one nucleolus, which stains with methylen green, as does 
the chromatic filament, and disappears in mitosis. Pisium , 
same cells : here there is one hat-shaped nucleolus, which stands 
in no connection with the “ Fadenknauel”; “ Sehr eigenthiim- 
lich ist das Verhalten des Nucleolus in den die Kerntheilung 
vorbereitenden Stadien. Anfanglich besteht derselbe aus 
homogener, stark lichtbrechender Substanz. Spater sind am 
Nucleolus eine dichte aussere und eine innere, bedeutend 
schwacher lichtbrechende Schichte unterscheidbar. Endlich 
findet man Stadien, auf denen neben dem noch unveranderten 
Fadenknauel ein sehr schwach lichtbrechender Korper gefunden 
wird, dessen Umrisse vollkommen demjenigen des urspriing- 
lichen Nucleolus entsprechen ”; finally even this disappears. 

Zacharias (’ 82 ) studied the epidermis cells of Phajus , and 
concludes that the nucleoli (one or two in number) consist of 
plastin. They do not dissolve in distilled water; swell with 
.if) nitric acid ; do not stain with methylen green; and 
dissolve in weak “ Kalilaugelosung.” 

Heuser (’ 84 ) studied the mitoses in the embryo sac of Fritil- 
laria imperialis. In the resting nucleus there are from five 
to nine nucleoli: “ Dieselben sind intensiv gefarbt und stehen 
in deutlich wahrnehmbarea Zusammenhang mit dem Nucleo- 
Hyaloplasma.” In the prophase of the mitosis they lose their 
staining power and apply themselves to the chromatin threads. 
He considers them, with Strasburger, “als Reserve-Behalter 
der Kernsubstanz ” (using the term “ Kernsubstanz ” as equiva¬ 
lent to “Chromatin”); their ground substance consists of 
plastin, permeated with chromatin. In Fritillaria , as well as 
in Galanthus and Leucojum , “ fliesst das gesammte Kernkor- 
perchen in die Kernelemente iiber, wahrend in anderen Fallen 
ein Ueberschuss an Plastin als ‘ Secretkorper ’ ausgeschieden 
werden mag.” 

Strasburger (’ 84 ), nuclei in the embryo sac of Fritillaria: in 
the spirem stage the large nucleoli disappear, “ wobei sich um 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


379 


dieselben der Kernsaft wieder zu farben beginnt.” He con¬ 
cludes that the nucleoli are not immediately taken up into the 
chromatic thread, but dissolve in the caryolymph; “ auch ist 
hiermit wohl sicher der Nachweis gegeben, dass sie nicht iden- 
tisch mit den Mikrosomen sein konnen.” The nucleoli arise 
in the meshes of the chromatin network. Strasburger agrees 
with Flemming that they represent a substance distinct from 
the chromatin and nuclear sap, but does not consider it to be 
a living substance, but rather a reserve stuff. 

Guignard (’ 85 ) investigated nuclear division in several species 
of plants. Lilium , young embryo sac: the nucleus usually 
contains a single nucleolus, which is very large, finely granular 
in structure, and situated excentrically between the strands of 
the chromatin network; with the double stain, methylen green 
and fuchsine, it stains red, while the chromatin stains green. 
At the time of the longitudinal division of the chromatin 
filament, the nucleolus commences to stain less intensely, 
vacuoles arise in it, and it finally fragments into small pieces 
which subsequently disappear; the fine granules appearing in 
the nuclear sap at this time are not derivatives of the nucleolus, 
but originate from the cytoplasm when the nuclear membrane 
vanishes. “ Dans le Lilium . . . rien ne fournit la preuve d’un 
apport direct effectue dans la formation du fuseau par le nucleole, 
dont la substance se dissout dans le sue nucl^olaire, pour 
s’incorporer et se melanger, . . . aux autres dements figures 
qui contiennent la chromatine.” In each daughter-nucleolus 
there are several nucleoli of unequal size; these disappear also 
in the subsequent mitosis. Clematis , embryo sac: the nucleoli 
in karyokinesis gradually decrease in size, and it seems “ comme 
si la plus grande partie de leur substance etait absorbee par les 
segments [chromatiques].” Northoscordum: here there are 
several large nucleoli which disappear when the spindle is 
produced, their substance being possibly incorporated in the 
chromosomes. In the metaphasic spirem they reappear in 
contact with the chromatin : “ leur aspect general fait supposer 
qu’ils naissent la ou on les aper$oit dans les jeunes noyaux . . . 
il est a croire que les nucleoles tirent une partie de leur 
substance, tout ou moins, du filament nucleaire auparavant 


38° 


MONTGOMERY. 


[Vol. XV. 


homog&ne. Ils se nourrissent ensuite dans le sue nucleaire. . . . 
Les nucleoles peuvent etre consideres comme une substance 
de reserve que se sdpare a un moment donne de la charpente 
nucleaire pour etre reprise par elle ulterieurement ”; he assumes 
that Strasburger’s “corpuscule du secretion” is a true nucleolus. 
“ Dans le Lilium et dans l’autres plantes, les noyaux filles 
n’offrent pas de nucleole avant d’entrer en division; en outre, 
leur aspect general au debut du phenomene est bien different 
de celui du noyau mere. ... Le fait qu’ils se separent du 
filament des que le noyau . . . arrive a l’etat de repos, pour 
etre repris par lui aux premiers stades de la division, permet 
de les consid^rer, avec M. Strasburger, comme une sorte de 
reserve.” 

Macfarlane (’ 85 ) studied nuclear division in Chara fragilis 
(fixation with osmic acid): the nucleus of the apical cell contains 
one nucleolus, in which lies an “ endonucleolus ” (a term here 
substituted for his earlier term “ nucleolo-nucleus ”). At the 
commencement of all cell divisions this part of the nucleolus 
first divides, then the nucleolus, last of all the nucleus. After 
this division of the apical cell a nodal and internodal cell 
are produced, and the former “continues to divide regularly, 
forming cells each with one nucleus and nucleolus. In the 
internodal complete cell division is henceforward absolutely 
arrested: but the earlier steps are taken ; for while the nodal 
cell has divided into three or four, the nucleolus of the inter¬ 
nodal has divided and redivided, so that four nucleoli are present 
in the nucleus of it. The internodal cell then increases rapidly 
in length, the four nucleoli meanwhile continuing to proliferate, 
so that in internodal cells, such as in the third removed from 
the apex, we soon get a large nucleus with many little dark 
nucleoli. The nucleus then divides in the simple manner 
figured by Johow, so that in the fourth internodal cell there 
may be two nuclei, each with many nucleoli, in the fifth, three 
or four nuclei, and so on, so that the internodal cells soon 
become multinuclear, and their nuclei multinucleolar.” The 
cortical nodal cells do not divide further, but “ their nucleoli 
follow the example of that of the internode . . . the consequence 
being that the cortical nodal, and soon after the cortical 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 381 


internodal cells, become multinucleolar ”; the nodal leaf cells 
proceed in the same way. From these observations Macfarlane 
concludes: “ in every active embryonic cell one nucleolus only 
is present in the resting state ”; in some cases a fluid globule 
is present in the nucleolus, and this probably represents a 
“ degradation of the endonucleolus.” “ The nucleolus, or more 
probably the nucleolo-nucleus, is the center of germinal activity, 
and that as we pass outwards to the periphery of the cell, this 
reproductive activity becomes less and less. . . . The result is 
that in all plants thus examined, after cell division has ceased, 
continued division of the cell contents from the endonucleolus 
outwards goes on. ... I venture, therefore, to regard it as a 
general principle that after cell formation has ceased, the cell 
contents (especially the endonucleolus and nucleolus) persist 
in their activity for a shorter or longer period; . . . the most 
exalted type of cell is one with abundant protoplasm containing 
a single nucleus, nucleolus, and endonucleolus; ... a cell with 
vacuolated protoplasm, one nucleus and two to four nucleoli 
is less exalted, while the multinuclear state is the most degraded 
form of cell.” 

Zacharias (’ 85 ) gives critical reviews of numerous preceding 
papers on nucleoli, besides observations of his own on various 
cells of plants. Galanthus nivalis , cells of the inner layer of 
the “ Fruchtknotenwand ”: the single nucleolus is about tV the 
size of the nucleus; examined in water it is homogeneous; after 
the action of absolute alcohol it appears to be composed of 
granules of various indices of refraction. Bast cells of Cucurbita 
pepo : the nucleoli, when stained with “ Blutlaugensalz-Eisen- 
chlorid,” become very intensely colored, while the remaining 
nuclear substance stains only faintly. In the cells of Spirogyra 
and of the asci of Lichens he finds that there are no “ nucleoles- 
noyaux,” such as Carnoy described. “ Alle Autoren stimmen 
gegenwartig darin iiberein, dass die Nucleolen bei der Kern- 
theilung verschwinden.” In opposition to Strasburger he 
contends that during the mitosis the dissolved nucleolar sub¬ 
stance might as probably enter into the formation of the spindle 
fibers as of the chromosomes. In Chara (observed living) each 
nucleus contains one large nucleolus, with vacuoles : “ Naht 


3 82 


MONTGOMERY. 


[Vol. XV. 


die Kerntheilung heran, so verliert der Nucleolus an Deutlich- 
keit, er erfahrt langsame Gestaltsveranderungen, die schliesslich 
einen amoboiden Charakter annehmen,” and the nucleolus 
gradually disappears (this process lasting a half hour); “i>£ 
Stunde spater wurden in jedem Tochterkern vier kleine Nude- 
olen bemerkt, nach y / 2 Stunden waren nur noch je zwei 
Nucleolen vorhanden und nach weiteren i y 2 Stunden nur noch 
je einer. . . . Bei der Verschmelzung bilden die Nucleolen 
zunachst einen bisquitformigen Korper, der sich dann spater 
kugelig abrundet. Die Deutlichkeit der Nucleolen nimmt 
wahrend des Vorganges der Verschmelzung stark ab, um spater 
wieder zu steigen.” Contrary to Strasburger and Tangl, he 
believes that no “ Paranucleolen ” wander out of the nucleus, 
but that where such have been observed, it has been due to the 
method of fixation. He notes that while the egg cells always 
contain nucleoli they are frequently absent in the male cells. 
“ In alternden Zellen sind Gestaltsveranderungen, Kleiner- 
werden und Schwinden des Nucleolus beobachtet worden. . . . 
Mir scheint es nicht begriindet zu sein, den Nucleolus als eine 
Ablagerung von Reservestoffen zu betrachten. Wesshalb 
sollte er nicht ein Organ der Zelle sein, wie es Flemming 
annimmt ? . . Against Strasburger’s views “ ist zu erwidern, 
dass wir fiber das active oder passive Verhalten der Nucleolen 
im ruhenden Zustande oder dem der Theilung fiberhaupt gar 
nichts wissen, und das Bestehen einer Organisation ffir die 
Nucleolen ebenso gut angenommen werden kann wie ffir irgend 
einen anderen Theil der Zelle.” 

Meunier (’86), Spirogyra: the single large nucleolus has a 
limiting membrane, and in the fresh state contains no vacuoles, 
vacuoles only appearing in the dying cell, and then are probably 
introduced drops of water. It stains with methylen green 
more intensely than any other structures of the nucleus, and 
also stains with acid picrocarmine, alkaline carmine, and haema- 
toxylin ; “ ainsi ... on constate que les matieres colorantes, 
reputees specifiques de la nucleine, limitent uniquement leur 
action efficace et significative au corps refringent et apparem- 
ment reticuld du nucldole.” After the action of nitric acid of 
from 2/0 to 4 Jo a reticulation is found in the nucleolus; a 10 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 383 

or I 2 j(> solution of the same acid dissolves this reticulation and 
only preserves the clear, non-refractive stroma ; 2 jfc or 4 jo 
hydrochloric acid solution shows the reticulation of the nucle¬ 
olus to be “ un boyau continu et pelotonne. ... Le filament 
chromatique du nucleole ne se digere pas dans la liquer digestive 
[sue gastrique]. . . . Nous ne craignons pas d’affirmer que 
le nucleole des Spirogyra reproduit fidelement, dans ses traits 
essentiels, la structure des noyaux les plus parfaits. II a une 
membrane propre, probablement une partie protoplasmatique, 
quoique fort r^duite ; il renferme toute la nucRine du noyau, et 
celle-ci est exclusivement confinde dans un dtui de plastine, 
qu’elle remplit plus ou moins complement. . . . Quoi qu’il 
en soit, nucleole par position, noyau par nature, on ne peut lui 
refuser le nom de nucleole-noyau, dans le sens attache a ce mot 
par J. B. Carnoy.” 

Schwarz (’ 87 ) studied the microchemistry of plant cells. He 
distinguishes the following substances in the nucleus : “ chro¬ 
matin,” “pyrenin” (nucleolar substance), amphipyrenin ” (sub¬ 
stance of the cell membrane), “ linin ” (achromatic fibrils), and 
“paralinin” (nuclear sap). The pyrenin and amphipyrenin 
“ stimmen in fast alien Reactionen iiberein, sie unterscheiden 
sich jedoch durch ihre Tingirbarkeit, indem das Pyrenin der 
Kernkorperchen Farbstoffe fast immer sehr leicht aufnimmt 
und festhalt, wahrend das Amphipyrenin nur wenig oder gar 
nicht tingirt wird. ... In den weitaus meisten Fallen liegt 
das Maximum des Nucleolusvolumens vor der Zone, in welcher 
der Kern sein Maximum erreicht, und in vielen Fallen tritt 
gerade dann die bedeutendste Verkleinerung des Nucleolus¬ 
volumens ein, wenn der Kern sein Volumen am starksten 
vergrossert. Es scheint mir demnach wahrscheinlich, dass ein 
Theil der Kernkorperchensubstanz direkt bei der Neubildung 
der iibrigen Kernsubstanz verbraucht wird.” 

Went (’ 87 ), mitosis in various cells of plants. Leucojum , 
embryo sac: at the commencement of the prophasis there are 
two large nucleoli, which lie between the fibers of the chromatin 
network ; later they become apposed to these fibers, and he 
notes how “die Masse des Nucleolus langsam in die des Kern- 
fadens ubergeht. ... Im Wandbelege des Embryosackes von 


384 


MONTGOMERY. 


[Vol. XV. 


Helleborus viridis scheinen die Nucleolen auch im Kernfaden 
aufgenommen zu werden”; and there is apparently the same 
process in Fritillaria imperialis. “ Bei den Kernen im Wand- 
belege des Embryosackes von Narcissus pseudonarcissus findet 
die Aufnahme des Nucleolus ungefahr wie bei Galanthus statt; 
er wird also von alien Seiten vom Kernfaden umwunden; 
allmahlich windet dieser sich wieder los. Oft ist dann der 
Nucleolus schon ganz aufgenommen, zuweilen aber werden 
noch Theile davon vom Kernfaden fortgeschleppt und bleiben 
dann wohl einmal sichtbar bis zum Anfang der Metaphase. 
Wenn man Praparate mit diamantfuchsin-jodgriin tingirt hat, 
sieht man, dass die Farbe des Kernfadens vor der Aufnahme 
des Nucleolus blaugriin ist, wahrend dieser letztere roth gefarbt 
ist; nach der Aufnahme des Nucleolus und wahrend der ganzen 
Meta- und Anaphase ist die Farbe des Fadens deutlich violett 
geworden, was naturgemass verursacht ist durch die Aufnahme 
des Nucleolus”; also during the mitosis of similar cells in 
Hyacinthus and Tulipa nucleolar substance is taken up by the 
nuclear filament. “ Ich glaube aus den hier mitgetheilten 
Thatsachen wohl den Schluss ziehen zu diirfen, dass in vielen 
Fallen wenigstens der Nucleolus beim Anfang der Kerntheilung 
im Kernfaden aufgenommen wird. . . . Am wahrscheinlichsten 
ist es wohl, dass, wo der Nucleolus vor der Theilung im Kern¬ 
faden aufgenommen wird, er sich nach der Theilung auch 
wieder daraus bildet.” 

Strasburger (’88) studied nuclear division in Spirogyra poly- 
taeniata. In the resting nucleus there is usually one large 
nucleolus, which disappears immediately before the formation 
of the nuclear filaments, and by dissolving in the nuclear sap 
causes the latter to stain more intensely : “ Als wahrscheinlich 
stellte ich [’ 84 ] es aber hin, dass die im Kernsaft geloste 
Nucleolussubstanz den Kernfaden als Nahrung diene. . . . Auf 
Grund meiner neueren Erfahrungen erscheint es mir iiberhaupt 
unwahrscheinlich, dass die Nucleolarsubstanz, auch nach ihrer 
Aufldsung im Kernsafte, den Kernfaden als Nahrung dienen 
sollte.” In each daughter-nucleus several nucleoli arise, and 
these have the same number, position, and size in the two nuclei; 
later the several nucleoli of each daughter-nucleus unite to form 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 385 

a single large nucleolus, and during this process the nuclear 
sap gradually loses its staining power. He shows that when 
the nucleolar substance is dissolved in the nuclear sap, and 
after the cell division, a portion of this substance plays a part 
in the production of the cellulose walls of the daughter-cells ; 
but he holds that not all of it is thus consumed, but that the 
nucleoli have probably some other, as yet unknown, function. 

Mann (’91) introduces a new method of differential nuclear 
staining: when plant tissues are stained for ten minutes in 
saturated solution of heliocin in 50 jo alcohol, and then from ten 
to fifteen minutes in a saturated aqueous solution of aniline blue, 
the nucleolus is red, the rest of the nucleus and the cell blue. 

Macfarlane (’92) constructs the following hypothesis, based 
on previous observations of his own and of Mann: “We would 
consider, then, that the nucleolus is the special chromatic and 
cell center ; that it sends out fine radiating processes — the 
intranuclear network — which partially fuse externally to con¬ 
stitute the nuclear membrane, the interspaces of the network 
being occupied by nucleoplasm concerned in metabolic change; 
that radiating continuations of the chromatic substance pass 
out beyond the nuclear membrane and form a network in the 
protoplasm, while we would suggest for future proof or disproof 
that they further may be continued through wall pores to form 
an intercellular chromatic connection. . . . We would thus 
view a plant as a group of connected hermaphrodite cells, . . . 
bound together by a fine chromatic ramification, in the center 
of which in each cell is the nucleolus.” 

Mann (’92) studied the cells of the embryo sac of Myosurus 
minimus. At the commencement of the conjugation of the two 
nuclei resulting in the formation of the primary endosperm 
nucleus, each nucleus contains “ a large deeply stained nucleolus 
enclosed by a very faintly stained nucleolar membrane,” and 
in each nucleus are also one or two smaller globules, which 
“ seem to originate thus : when the nuclei about to conjugate 
have come in contact, one or two small nucleoli arise by the 
unequal division of the primary nucleolus. . . . These secondary 
nucleoli seem to have at first the power of division, but 
gradually they lose this power and their property of becoming 


3 86 


MONTGOMERY ,. 


[Vol. XV. 


deeply stained, and change into globular colloid-looking masses 
with a central more deeply stained spot. I propose to call these 
bodies paranucleoli, because of their origin they may always be 
found in the micropylar nucleus and occasionally also in the 
antipodal nucleus.” When these nuclei begin to conjugate, 
the large nucleoli of both fuse to form the single nucleolus of 
the primary endosperm nucleus ; at the same time a new struc¬ 
ture makes its appearance, in close contact with the nuclear 
membrane of the primary endosperm nucleus : “ This body . . . 
corresponds, I believe, to the nucleolar membrane of the 
antipodal nucleus” ; it; is at first granular, later homogeneous. 
Still other, smaller spherical bodies later appear in the nucleus, 
which may have some connection with the paranucleoli. Finer 
structure of the nucleolus: in the nucleolar membrane “ a 
number of very minute dark radially placed pores or striae can 
be observed, and . . . these striae are continued into very 
delicate cilia-like fibrils radiating out from the nucleolar mem¬ 
brane into the nuclear hyaloplasm. . . . The nucleolus is 
differentiated into an outer zone and an inner zone. The outer 
zone is less deeply stained, and on careful examination is found 
to be made up of a circle of peripheral endonucleoli, which are 
slightly elongated radially. The inner zone of the nucleolus 
is very darkly stained, and shows a number of large and irregu¬ 
larly disposed endonucleoli.” The structure of the nucleolus 
may be somewhat different in other stages of its development, 
thus it may be composed of “ (i) A thin unstained nucleolar 
membrane; (2) a great number of peripheral endonucleoli; (3) 
a deeply stained, apparently structureless, layer; (4) a corona 
of minute, slightly elongated, endonucleoli surrounding (5) a 
large central endonucleolus. ... In a resting cell, . . . the 
center of the nucleolus is occupied by a large endonucleolus, 
which sends out minute fibrils through the nucleolar sub¬ 
stance. ... I believe the endonucleolar fibrils probably to pass 
through the finer pores in the nuclear membrane”; and Mann 
conjectures that ‘‘the endonucleolar filaments constitute the 
linin element of the chromosomes.” Functions of the nucle¬ 
olus: it is “concerned in the assimilation of food-material.” 
He holds “the nuclear chromatin to be less highly elaborated 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 387 

and less assimilative albuminoid material than the nucleolar 
chromatin. On the assumption just stated, we could explain 
also why we find ... at the time of maturation portions of 
nucleolar matter detaching themselves from the main nucleolus 
to undergo a peculiar gelatinous change. The gelatinous change 
would correspond to a conversion of the assimilative material 
into achromatic elements, an explanation which would also 
explain the disappearance of nucleoli during the division of a 
cell. ... I believe the hypothesis that the nuclear chromatin- 
segments and perhaps the nucleoli are organs for the conversion 
of assimilated material into material directly available for the 
achromatic elements of the cell to be not quite erroneous.” In 
the mechanism of cell conjugation: “The endonucleolar fibers 
running through the body-plasm of the two sexual cells . . . are 
brought into contact with one another whenever the pseudopo- 
dial processes of the two cells have met. As soon as an union 
of fibrils has taken place, each fibril will commence to contract 
similarly to a muscular fibril,” which results in drawing the two 
nuclei, afterwards also the two nucleoli, together; thus the 
endonucleolus is the “ tropic center ” of the cell. 

Rosen (’92a) studied the differential staining of the nuclear 
elements in plants. Flowers of Scilla: in the nuclei of the 
“ Biindelparenchym ” are numerous large nucleoli, which differ 
in form and size; the one or two larger ones, “ Eunucleoli,” are 
each surrounded by a clear space, but none is present around 
the smaller “ Pseudonucleolen.” With the double stain, Alt- 
mann’s acid fuchsine and methylen blue, the Eunucleoli stain 
red and the Pseudonucleoli blue, or vice versa. Similar cells 
of Hyacinthus: by the application of the double stain, aqueous 
solutions of fuchsine and methylen blue respectively, the 
Eunucleoli stain red, the Pseudonucleoli blue ; but when these 
stains are applied in the reverse order, the nucleoli stain 
reversely. He considers, following Auerbach (’90), that the 
Eunucleolus is erythrophilic, the Pseudonucleoli kyanophilic, 
the latter staining as does the chromatin network. “Meine 
Pseudonucleolen aber sind eben offenbar weiter nichts, als 
besonders selbstandig ausgebildete Bestandtheile des chroma- 
tischen Kerngeriistes und sind wie dieses und sein Produkt, der 


MONTGOMERY. 


[Vol. XV. 


388 

Kernfaden, kyanophil”; these disappear before the mitosis, 
while the Eunucleoli remain until about the end of the spirem 
stage. Vacuoles arise only in the Eunucleoli. 

Rosen in a second paper (’92b) presents further observations 
upon nucleoli. Myxomycetes: the spore nucleus contains one 
large nucleolus. Fuligo septa , plasmodium : one large, cyano- 
philic nucleolus, which he terms “ Mittelkorperchen,” since in 
the atypical mitosis this body lies in the middle of the pole 
plate, and disappears at the end of the nuclear division. Syn- 
chrytrium: one large nucleolus with several vacuoles ; in the 
first mitosis the division of this nucleolus precedes that of 
the nucleus, but during subsequent divisions the nucleoli 
vanish. In Cystopus there is no nucleolus. 

Schottlander (’92), cells of cryptogams : the nucleus consists of 
a blue-staining substance (network), and a red-staining (nuclear 
membrane, nucleoli). Egg cell of Gymnogramme chrysophylla : 
here are one or several large nucleoli, each surrounded by a 
vacuole; in the ripe egg the nucleoli are filled with small glob¬ 
ules. Egg cell of Chara: the nucleoli contain vacuoles, which 
later become so large in the largest nucleoli that they become 
polygonally flattened against one another, and their thin walls 
then present the appearance of a network within the nucleolus. 

Demoor (’93), mitosis of Tradescantia: the nucleoli gradually 
disappear during the prophase. 

Gjurasin (’93) investigated the nuclear division of Peziza. 
In the nucleus is one large, excentric nucleolus, which stains 
red with Flemming’s triple stain, while in it as many as six 
granules may occur, and these stain violet. In the mitosis 
these granules disappear, but otherwise the nucleolus does not 
change at first, but occupies its original position within the cell, 
though now in the cytoplasm; eventually it disappears gradually. 
In each daughter-nucleus a new nucleolus arises, which appar¬ 
ently has no genetic connection with the mother-nucleolus (now 
vanished). “ Ich bin der Ansicht, dass . . . das Kernkorper- 
chen nicht eine Art von Reservestoff darstellt, sondern ein 
specifisches Organ des Zellkernes ist.” 

Karsten (’93), nuclear division of Psilotum : in the resting 
nucleus are two or three nucleoli, which are homogeneous, oval 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 389 

or spherical, and after haematoxylin-eosin, stain a rose color, 
while the chromatin is blue. At the time of the appearance of 
the chromosomes, “ treten die Nucleolen aus den sich zusam- 
menordnenden Plasma und lassen sich hier in Form scharf 
umschriebener, homogener, roth gefarbter Kiigelchen nach- 
weisen.” Usually two nucleoli wander out, at least never more 
than two were found outside of the nucleus. These two come 
to lie at opposite poles of the nucleus, occupying the positions 
of centrosomes; and when the longitudinal splitting of the 
chromosomes takes place, each of the nucleoli also divides into 
two. Karsten believes these nucleoli are identical with the 
centrosomes of Guignard; but he does not explain what becomes 
of the third nucleolus during the division. 

Lauterborn (’93), quoted by Karsten (’93), diatoms : there 
is a centrosome lying in a concavity of the nucleus; he noticed, 
further, “ beim Beginn der Theilung aber zwischen Kern und 
Centrosom noch ein anderes Gebilde —, welches im spateren 
Verlauf der Karyokinese eine sehr bedeutsame Rolle spielt, 
namlich die Anlage der Centralspindel ”; this body must be 
derived either from the nucleus or the centrosome (I mention 
it here since it may in the future be found to have some con¬ 
nection with a nucleolus). 

Moll (’93) studied karyokinesis in Spirogyra. There are one 
or two nucleoli, which stain more intensely with gentian violet 
than any other portion of the nucleus. They may be vacuolar 
in structure, or contain a skein of chromatin ; they appear 
homogeneous only when too deeply stained. The skein struc¬ 
ture (the skein itself staining as chromatin) is found in resting 
nuclei, as well as in the prophases of mitosis, and at the same 
time vacuoles may be present. He assumes that the thread in 
the nucleolus contains all the chromatin of the resting nucleus, 
and “ that by the nucleolus the chromatin substance for the 
segments [chromosomes] is furnished”; this chromatin leaves 
the nucleolus in mitosis, and “ it seems as if the chromatic sub¬ 
stance were squeezed from the nucleolus by an aperture.” After 
the chromatin skein has left the nucleolus, the latter disappears. 

(Strasburger’s paper, ’93, was reviewed under the head of 
zoological literature.) 


390 


MONTGOMERY. 


[Vol. XV. 


Wager (’93), nuclear division in Hymenomycetes agaricus: 
each nucleus of a basidium contains one large nucleolus, besides 
the nuclear network. The two nuclei of the basidium fuse 
together and form one nucleus, in which afterwards the two 
nucleoli later fuse to form one nucleolus. This latter is 
often vesicular in structure. In the mitosis it lies close to the 
nuclear membrane, it gradually loses its staining intensity, 
decreases in size, and finally disappears; at the same time the 
cytoplasm in its neighborhood stains more deeply. But some¬ 
times it persists until the diaster stage. “ From the fact that 
the chromosomes begin to stain red at the time of the disappear¬ 
ance of the nucleoli, it would further appear that the former 
can take up nucleolar substance from the nuclear sap, and as 
fast as the nucleoli disappear the chromatic elements become 
more deeply stained red.” In A. stercorarius , in the daughter- 
nucleus, “the chromatin mass appears to be transformed at 
once into the nucleolus,” and only later a chromatin network 
appears. “ I would suggest that the nuclear threads take up 
the dissolved nucleolar substance at some period during the 
division, and carry it over into the daughter-nuclei, to be given 
up again later as the nucleoli of the latter. . . . But a certain 
quantity of the dissolved nucleolar substance probably escapes 
into the cytoplasm when the nuclear membrane disappears, and 
this would be taken up at a later stage into the daughter-nuclei, 
as is shown by the increase in size of the nucleoli, and by the 
decrease in the capacity of the protoplasm for taking up stains.” 

Zacharias (’93) finds in plants that the nucleolus and cyto¬ 
plasm are erythrophilic, the nuclein (chromatin) network is 
cyanophilic. 

Belajeff (’94), “ Pollenmutterzellen ” of Larix: after the 
disappearance of the nuclear membrane in mitosis the nucleolus 
becomes gradually smaller and then disappears; several nucleoli 
reappear within each of the daughter-nuclei. “ Es ist zu 
bemerken, dass nach der Auflosung der Nucleolen der Mutter- 
zelle im Zellplasma eine gewisse Anzahl grober Kornchen 
erscheint, welche mit Safranin farbbar sind. Mit dem Beginn 
der Nucleolenbildung in den Tochterzellen verschwinden die 
Kornchen vollkommen. . . . Ich erklarte mir die Ergebnisse 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


391 


meiner Beobachtungen derart, als losten sich die Nucleolen, nach 
vorausgegangener Auflosung der Kernmembran, unter der Ein- 
wirkung der in die Kernhohle aus dem Zellplasma gedrungener 
Substanzen, ganzlich auf, um spater durch den Einfluss des 
Kernsaftes, der die ganze Zelle durchdrungen, wieder hergestellt 
zu werden, indem der Kernsaft die Nucleolensubstanz im Zell¬ 
plasma so zu sagen gerinnen macht. Nach der Bildung der 
Tochterkerne, welche ihren Kernsaft aus dem Zellplasma 
absorbiren, werden die Kornchen abermals vom Zellplasma auf- 
gelost, um zum zweitenmal im Inneren der jungen Kerne 
(Tochterkerne) in der Gestalt von Nucleolen zu erscheinen.” 
In Fritillaria and Lilium also the nucleolus is dissolved after 
the disappearance of the nuclear membrane. 

Humphrey (’94) studied the “ Pollen-” and “ Sporenmutter- 
zellen ” of Convalleria , Ceratozamia, Osmunda , and Psilotum , 
and cells from the apex of the root of Vicia and Hyacinthus. 
The nucleolar substance is usually not to be found in the 
cytoplasm during mitosis. The nucleoli are “keine indivi- 
duellen Bestandtheile, sondern unbestimmte Massen von Nucle- 
olarsubstanz, und ihr Vorkommen im Cytoplasma hat keine 
weitere Bedeutung als zu zeigen, dass eine Communication 
zwischen Kernhohle und Cytoplasma bisweilen, wenn auch 
nicht immer, sich herstellen kann und dass entweder die Nucle¬ 
olen in einigen Fallen aus der Kernhohle, bevor sie von den 
karyokinetischen Kraften angegriffen werden, austreten konnen, 
oder dass die Menge der Nucleolarsubstanz in einem Kerne 
grosser sein kann, als diese Krafte zu losen oder zu verbreiten 
vermogen. . . . Die ‘Vacuolen’ der Nucleolen scheinen mir 
das naturliche Resultat der nachherigen T rennung der fliissigeren 
von den festeren Theilen der Nucleolarsubstanz zu sein. . . . 
Wenn also Zimmermann [’93] den Satz aufstellt ‘ Omnis 
nucleolus e nucleolo/ so kommt er zu einer Verallgemeinerung, 
die nicht zulassig und derjenigen ‘Omnis nucleus e nucleo ’ 
nicht gleichwerthig ist.” In every nucleus of the “ Pollensacke ” 
of Ceratozamia there is a large, peripherally placed paranucle- 
olus (Strasburger): “ In extremen Fallen kann die Anhaufung 
von Substanz eine so grosse sein, dass die Kernmembran 
hier bedeutend hinausgestossen wird. . . . Auf der Fuchsin- 


392 


MONTGOMERY. 


[Vol. XV. 


Jodgrun tingirten Schnitten werden die Paranucleolen weder 
reinroth wie die Nucleolen, noch blaugriin wie die chromatische 
Substanz gefarbt, vielmehr nehmen sie eine Zwischennuance, 
welche mehr der des Chromatins als der der Nucleolen ahnelt, 
an”; he believes these paranucleoli to be artefacts. In con¬ 
tradiction to Karsten (-93) he found no body in Psilotum 
comparable to a Nucleo-Centrosoma. 

Zacharias (’94) concludes, from numerous observations on 
cells of plants that as the size of the nucleus increases (or 
decreases) with the size of the cell, so also that of the nucleolus 
increases (or decreases) with the size of the nucleus. 

In Rosen’s (’95) contribution a large number of new facts 
are recorded, which may be briefly mentioned. The kyanophilic 
nucleoli of Auerbach “ sind eben keine Nucleolen und bediirfen 
als wenig constante Theile des Chromatingeriistes iiberhaupt 
keines besonderen Namens.” Hyacinthus: in meristem nuclei 
all the nucleoli except the smallest lie in special clear spaces, 
and though fibrils are rarely found in connection with them, 
“gleichwohl muss das Kernkorperchen in seiner scheinbar 
schwebenden Lage wohlbefestigt sein, da es . . . stets seine 
Lage im Centrum seines Hofes bewahrt.” The large nucleoli 
of the “ Gefasszellen ” become vacuolar as they increase in size. 
In mitosis of root cells the nucleoli become gradually dissolved 
within the nucleus in some species, in others they are extruded 
into the cytoplasm; in the latter cases “ erfolgte die Zerkliiftung 
und Auflosung des Nucleolus viel langsamer, sodass bei dem 
Schwinden der Kernmembran noch bedeutende Nucleolarreste 
vorhanden waren.” The nucleoli reappear in the dispirem 
stage before the daughter-nuclei have produced membranes, 
and the new nucleoli stain from the commencement intensely; 
from which the general conclusions are drawn : in the prophase 
the diminishing nucleolar substance penetrates, perhaps as a 
micellar solution, into the cytoplasm, and this process may 
cease before the nuclear membrane has disappeared. In some 
cases larger particles of nucleolar substance may penetrate 
into the cytoplasm, but only after the nuclear membrane has 
disappeared, and these particles become subsequently dissolved 
in the cytoplasm; in either case “ das losende Agens muss wohl 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


393 


der Kernsaft sein, vielleicht unter Mitwirkung eines nur 
wahrend der Prophasen gebildeten Enzyms. Wahrend der 
Anaphasen wandert die Nucleolarlosung als solche in den Raum 
der Tochterkerne ein, und hier wird die Nucleolarmasse wieder 
fest. Bei der Hyacinthe — und anderen Objekten — erfolgt 
die Rekonstituirung der Nucleolen auch ausserhalb der Dis- 
piremfigur. Die derart im Cytoplasma entstaiidenen Nucleolen 
wandern, wie ich glauben mochte, in die Tochterkerne ein, ehe 
sich diese mit einer Kernmembran umhullen; wenn letzteres 
geschehen ist, so findet man anscheinend niemals mehr Nucle¬ 
olen im Cytoplasma, die, wenn iiberhaupt, auch wohl nur nach 
nochmaliger Auflosung in den Kernraum gelangen konnten. 
Nicht ganz unmoglich scheint es mir, dass die Nucleolen, die 
man an fixirten Praparaten . . . im Cytoplasma auffindet, doch 
durch die coagulirende Wirkung des Fixirungsmittels ent- 
standen sind. Ich glaube aber, dass dies von keiner grossen 
Bedeutung ist, denn an den Stellen, wo wir extranucleare 
Nucleolen vorfinden, muss dann die Masse der Kernkorperchen 
als Losung angesammelt gewesen sein.” Also in the mitosis 
of root cells of Aspidistra , are nucleolar fragments seen in the 
achromatic spindle. Root cells of Phaseolus: in the resting 
stage there is a single nucleolus; in the mitotic prophase 
it becomes first lobular, then lengthened in the direction of 
the spindle, while at the same time it is undergoing a slow 
dissolution; “ wenn die Spindel gebildet und die Kernwandung 
verschwunden ist, sieht man fast stets inmitten der zur Kern- 
platte angeordneten Chromosomen einen mehr oder minder 
ansehnlichen Nucleolarrest, welcher in derselben Richtung wie 
die Chromosomen und die Spindelfaden gestreckt ist. Dieser 
Nucleolarrest wird nun in der Mitte eingeschniirt, sodass er 
Hantelform erhalt; die beiden Halften reissen schliesslich von 
einander und gelangen an die Spindelpole. In anderen Kernen 
wird der Nucleolarrest einseitig aus der Kernplatte herausge- 
drangt oder auch doppelt getheilt; endlich findet sich meist an 
einem oder an beiden Spindelpolen ein Restchen des Nucleolus; 
seltener liegt ein solches neben der Spindel. Die Auflosung 
ist nun meist bald beendigt”; and only exceptionally is there 
a minute nucleolar remnant in the cytoplasm at the end of 


394 


MONTGOMERY. 


[Vol. XV. 


mitosis. “ Unzweifelhaft sind auch bei Phaseolus multiflorus 
die Nucleolen der Tochterkerne Neubildungen. Wenn auch 
die Nucleolarsubstanz moglicherweise bei der Karyolyse er- 
halten bleibt und sich in den Tochterkernen nur wieder auf 
Neue sammelt, so besteht doch keine von Generation zu 
Generation sich fort spinnende Continuitat in den Nucleolen 
' als solchen und von einem * omnis nucleolus e nucleolo * 
[Zimmermann] kann keine Rede sein.” Root cells of Vicia 
faba: the nucleolar mass diminishes as the cell degenerates; 
“ dieselbe stellt das erste Zeichen der Kerndegeneration . . . 
dar und ist, wie sonst, mit einer Zertheilung des Nucleolus 
verbunden,” while a large nucleolus surrounded by a clear 
space is an embryonic condition. In the mitosis of these cells 
no nucleolar fragments pass into the cytoplasm, and in each 
daughter-nucleus two nucleoli arise which subsequently fuse 
into one. In opposition to Lavdowsky (’94), he contends that 
the centrosomes have no genetic connection with nucleoli, and 
that the nucleolar substance does not serve as nourishment for 
the chromosomes ; “ nichtsdestoweniger ware es voreilig zu 
behaupten, dass von der Substanz der Nucleolen nichts in die 
Fadensegmente gelangen konne. . . . Die Violettfarbung der 
Segmente in den spateren Phasen der Karyokinese . . . konnte 
auf eine Einlagerung erythrophiler Nucleolarsubstanz in den 
kyanophilen Kernfaden schliessen lassen.” In buds of Psilo - 
turn triquetrum the nucleoli are excentric, while in most plants 
they have a central position. In the mitosis nucleolar frag¬ 
ments are extruded into the cytoplasm (in agreement with 
Zimmermann, in opposition to Karsten and Humphrey), and 
none of the extruded masses can be regarded as centrosomes 
(against the view of Karsten). Three nucleoli usually arise in 
each daughter-nucleus : “ Sie entstehen nahe der Peripherie 
des j ungen Kerns, oft in Contakt mit dem Cytoplasma, bevor 
die Tochterkerne sich mit einer Membran umschliessen und 
verschmelzen spater nicht miteinander.” In the mitosis of 
sporangia the nucleoli are usually “ aus den karyokinetischen 
Figuren ausgestossen ” ; and the “ Secretkorperchen ” of 
Strasburger is a true extruded nucleolus. 

Strasburger (’95, cited by Lauterborn, Zool. Centralbl.y 1896 ) 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


395 

concludes that the nucleolar substance, dissolved in the nuclear 
sap, may be used in the production of the spindle fibers. 

Koernicke (’96), study of mitosis on Triticum: in the devel¬ 
opment of the embryo sac when the two pole nuclei fuse 
together, the two nucleoli also join to form one. In the mitosis 
of the pollen the nucleolus always disappears before the forma¬ 
tion of the spindle, but it could not be determined whether it 
takes any part in the formation of the latter. 

Lauterborn (’96), nuclei of diatoms: there are several nucleoli 
present; in the spirem stage of division they commence to 
gradually disappear ; “ es scheint mir ziemlich sicher, dass ihre 
Substanz mit derjenigen der Chromatinkornchen und des 
Liningeriistes zur Bildung der Knauelfaden verbraucht wird.” 
It is important to note that the central spindle arises outside 
of the nucleus, before the nucleoli begin to disappear, so that 
there can be no genetic connection between the two. 

Poirault and Raciborski (’96), binucleated (“conjugate”) 
Uredineae during the production of the ascidiospore generation : 
in the mitosis the nucleolus becomes extruded into the cyto¬ 
plasm, almost always in the equatorial plane. “ Bei manchen 
Arten bleiben sie sehr lange erhalten so z. B. bei Peridermium 
Pini acicola , wo neben den langst ruhenden, mit neuen Nukleo- 
len versehenen Kernen noch in den Plasma, die alten Kern- 
korperchen der Elternkerne herumirren. Mit den Centrosomen 
haben somit diese extranukleolaren, vakuolirten Nukleolen 
nichts zu thun.” 

Zimmermann (’96), a general critical summary upon the 
vegetable nucleolus, with consideration of a part of the previous 
literature. Nucleoli are almost always present in the cells of 
the higher plants, and are of wide occurrence also in the lower 
forms ; double staining serves to differentiate them from the 
chromatin. There are usually from one to three to a nucleus, 
but in the embryo sac of Lilium martagnon there are from 
twenty to thirty. In Chara the older nuclei show the nucleolar 
substance in the form of very numerous, irregular fragments. 
The distinction of “ Hauptnucleolus ” and “ Nebennucleolus ” 
is not tenable, since the latter may be possibly chromatin 
globules. “Mit dem Chromatingeriist scheinen die Nukleolen 


39 6 


MONTGOMERY. 


[Vol. XV. 


innerhalb der ruhenden Kerne in keinem Falle in direkter 
Verbindung zu stehen.” The space frequently observed around 
the nucleolus is probably not an artefact. Its substance is 
probably homogeneous ; “ als die alleinigen mit Sicherheit 
nachgewiesenen Einschlusse derselben konnen Vakuolen ange- 
fiihrt werden. . . . Diese Vakuolen sind dem gewohnlichen 
Einschluss in Kanadabalsam haufig ganz oder teilweise mit 
Luft erfullt oder stellen luftleere Raume dar. Sie erscheinen 
dann bei hoherer Einstellung schwarz, bei niederer etwas rot- 
lich, und es diirften wohl die namentlich in der die Kerne 
beilaufig behandelnden Litteratur vorliegenden Angaben liber 
stark lichtbrechende Einschlusse der Nukleolen zum Teil auf 
derartige Bilder zuriickzufiihren sein ” (eg, the “ endonucleoli ” 
described by Mann). During mitosis nucleolar bodies are 
often found in the cytoplasm, and such are probably extruded 
nucleolar fragments ; “ immerhin muss aber die allgemeine 
Giiltigkeit des friiher von mir als moglich hingestellten Satzes 
omnis nucleolus e nucleolo nach den neueren Untersuchungen 
als nicht sehr wahrscheinlich angesehen werden.” In the Pol- 
lenmutterzellen of Lilium martagnon the nucleoli “ zerfallen 
. . . in sehr zahlreiche kleine Kugeln, die . . . im Aster- 
stadium ungefahr gleichmassig liber den gesammten Zellinhalt 
zerstreut sind.” He made similar observations also on Hya- 
cinthus candicans, Fritillaria imperialism young sporangia of 
Equisetum and Psilotum, cells of the root apex of Vida, and 
stem apex of Phaseolus and Psilotum. There is also an extru¬ 
sion of nucleolar substance in Chara , but it is doubtful whether 
this process occurs in other low forms. This extruded sub¬ 
stance may in some cases, but perhaps not as a rule, return 
into the daughter-nuclei. That in mitosis the nucleolar sub¬ 
stance may be incorporated into the chromosomes, “ sei noch 
erwahnt, dass ich neuerdings an den Kernteilungsfiguren des 
Embryosack-Wandbelags von Lilium martagnon nach der 
Fixierung mit Chromsaure und Platinchlorid und Farbung mit 
Fuchsin und Jodgriin in den Endstadien des Spirems beobachten 
konnte, dass einzelne rote Kugeln, die ausserdem auch in 
grosser Zahl in der Umgebung der betreffenden Kerne zu 
beobachten waren, den violettgefarbten Chromosomen teils 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


seitlich ansassen, teils auch ganz von denselben aufgenommen 
waren, so dass sie . . . kleine Auftreibungen an denselben 
bildeten.” It is doubtful whether the nucleoli have any genetic 
connection with either the centrosome or the nuclear membrane. 
In the synapsis (Moore, ’95) of the nucleus the nucleolus 
becomes flattened against the nuclear membrane in most 
Angiospermia , having thus on section a sickle shape (“ Sichel- 
stadium ”) ; and the coincidence of this form of the nucleolus 
with the synaptic stage “macht es jedenfalls sehr wahrschein- 
lich, dass die im Sichelstadium eintretenden Metamorphosen 
den Nukleolus eine gewisse Bedeutung besitzen.” 

Debski (’97), Chcira: the space surrounding the large nucleo¬ 
lus is caused by shrinkage of the latter, due to the fixing fluids, 
and is not present in life. In the nucleolus are numerous 
vacuoles which may become confluent. Within the cytoplasm 
occur extranuclear nucleoli, which stain like the others. In 
the mitotic prophase the nucleolus usually divides into two, 
and the latter either gradually diminish in size and finally dis¬ 
appear or else they persist for a while after the disappearance 
of the nuclear membrane. Then the extranuclear nucleoli 
collect at the poles of the spindle and “ bewegen sich wahrend 
der Metakinese von beiden Seiten her gegen den Ort der 
spateren Zellplattenbildung und verschmelzen dabei nicht 
seiten wahrend des Diasterstadiums miteinander zu unregel- 
massigen Kugeln, Klumpen und Faden ... die nucleolenar- 
tigen Korper sind spater, nach der Bildung der Zellplatte und 
der Membran, nicht mehr dort zu sehen ; es finden sich alsdann 
nur noch wenige durch das ganze Plasma der Zelle zerstreut, 
oder sie fehlen, besonders in den alteren Zellen ganzlich. 
Einige, wahrscheinlich solche, welche wahrend des Diasters 
nicht in die Zellplattenebene geriickt sind, finden sich wahrend 
des Dispirems in der Nahe der Tochterkerne ein ; spater sind 
sie zwischen den Faden des Kerngerustes zu sehen ; in spateren 
Stadien findet man an ihrer Stelle einige kleine Nucleolen, 
deren Zahl immer mehr beschrankt wird, so dass sich schliess- 
lich gewohnlich in jedem Kern ein einziger grosser Nucleolus 
befindet.” 

Fairchild (’97), Basidiobolus: “Das Verschwinden des 


398 


MONTGOMERY. 


[VOL. XV. 


Kernkorperchens . . . spricht entschieden fur Strasburgers 
Annahme, dass es zur Bildung der Spindelfasern benutzt 
werde.” 

Harper (’97), ascus of Erysiphe: the nucleolus and the cen- 
trosphere stain in the same way, and “ die achromatischen 
Fasern, aus welchen diese intranuclearen Strahlenkegel gebildet 
werden, entstehen wahrscheinlich grosstentheils auf Kosten 
der Kernkorperchensubstanz, die zu dieser Zeit regelmassig 
verschwindet.” 

Huie (’97), cells of Drosera: the nucleoli (“ nucleolar chromo¬ 
somes”) are spherical and usually central; “endonucleoli” 
are enclosed spaces, not granules. During the process of food 
assimilation by the nucleus the nucleolus becomes smaller, and 
its vacuoles less apparent. 

Lidforss (’97) gives a thorough review of the “ Sichelstadium ” 
(Strasburger’s “ Sekretkorperchen ”) of the nucleolus in plant 
cells, as also the results of observations of his own on the 
embryo sac. Tulipa: at first there are several small nucleoli 
within the nuclear cavity, which later by their fusion produce 
a large one which becomes flattened against the nuclear mem¬ 
brane (the process is essentially the same in Fritillaria, Anthe- 
ricum , and Lilium). Gagea : the nucleolar changes are as in 
the preceding forms, except that when the nucleolus reaches 
the periphery it remains spherical; this is also the case in 
Ornithogalum. Oenothera: in the youngest cells there is one 
central nucleolus ; subsequently this flattens against the nuclear 
membrane, but finally wanders back to the center and becomes 
spherical. He concludes that in the angiosperms the sickle 
stage of the nucleolus is a normal phenomenon, as is also its 
excentric position. In male and female germ cells these meta¬ 
morphoses occur at corresponding stages, namely, when the 
reduction of the chromatin takes place ; “ indessen bleiben 
vorlaufig alle Speculationen iiber die Bedeutung des Sichel- 
stadiums von problematischen Werth. . . .” 

Mottier (’97), cells of Podophyllum and Lilium: in mitosis, 
at the time of disappearance of the nuclear membrane, the 
nucleolus breaks into fragments of various size. “ Bei der 
Anlage der vielpoligen Spindel nun treten im Cytoplasma 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 399 

kleinere, dem Nucleolus ahnlich tingirte Korper auf. ... Es 
unterliegt keinem Zweifel, dass dieses die zerfallene Kernkor- 
perchensubstanz darstellt . . . nachdem der Tochterkern mit 
einer Wandung versehen wurde und in ihm Kernkorperchen 
zum Vorschein kamen, sind oft noch extranucleare Nucleolen in 
dem Cytoplasma zu sehen. Dasselbe gilt fur die zweite 
Theilung. Ob die in dem Tochterkern zum Vorschein kom- 
menden Kernkorperchen aus den im Cytoplasma liegenden 
Korperchen entstehen, lasst sich nicht feststellen. Hingegen 
ware hier hervorzuheben, dass die im Kern wieder entstehenden 
Kernkorperchen stets in Contakt mit den Kernfaden sich befin- 
den . . . meine Ansicht geht aber dahin, dass in den Kern¬ 
korperchen ein Kraftvorrath gegeben ist, welcher der Zelle 
nach Bedarf zur Verfiigung steht.” 

Pennington (’97), cells of Spirogyra treated with . 1478/0 
palladious chloride : “ The nucleolus showed a dark bounding 
layer of double contour. *. . . The dark layer is undoubtedly 
a true membrane dividing the nucleolus from the nucleus.” 

Strasburger (’97) reiterates his view (’95) that in plant 
mitoses the achromatic spindle is formed from nucleolar sub¬ 
stance, and that also the “Zellplatte” and “ Centralspindel- 
korperchen” of animal cells must be of nucleolar origin. 

Swingle (’97), algae ( Sphacelariaceae) : the vacuolization of 
the nucleoli occurs simultaneously with the separation of the 
two centrosomes, and probably at the same time that the differ¬ 
entiation of the chromosomes occurs. Though “die schnelle 
und vollstandige Auflosung der iibrigen Substanz des stark 
vacuolisirten Kernkorperchens findet statt, wenn die Spindel- 
fasern an den Polen einzutreten beginnen,” there yet seems to 
be no direct proof that these fibers have their origin in nucleolar 
substance. “ Konnte er [Nucleolus] nicht eher einen speciellen 
Vorrath organischer Nahrung zur Erhaltung des Kinoplasmas 
wahrend der Karyokinese vorstellen ? ” 

C . Synonyms of the Term Nucleolus. 

Since there are quite a large number of synonyms of the 
nucleolus, they may for convenience’ sake be classified together 


400 


MONTGOMERY. 


[Vol. XV. 


at this place. Certain of the following terms, however, apply 
not to the true nucleoli but to the Caryosomata. 

German writers .—Nucleolus (Valentin) Keimfleck, Keimkern, 
macula germinativa (Wagner); Kernkorper (chen) (Schwann, Val¬ 
entin); Keimkorper(chen); Wagner’scher Fleck; Binnenkorper 
(Rhumbler); Hauptnucleolus, Nebennucleolus (Flemming); 
Metanucleolus (Hacker) ; Plasmosoma (Ogata); Formations- 
nucleolus (Marshall); Kernfleck, Nucleolide, Morulit (Frenzel); 
Nucleolo-Centrosoma (Keuten); Mittelkorperchen, Eunucleolus 
(Rosen); Nucleolkorperchen (Lonnberg); Stammnucleolus, 
Nebenkiigelchen (Auerbach); Hauptkeimfleck, Nebenkeimfleck 
(Leydig) ; Chromatin-Nucleolus, Paranucleolus (R. Hertwig). 

English and American writers. — Wagnerian vesicle, ento- 
blast (Agassiz); pronucleolus (Mark); nucleole, germinal spot, 
germinal dot, principal nucleolus, accessory nucleolus, proto- 
macrosome (Greenwood). 

French writers. —Nucleole, tache germinative; pseudonucleole 
(Van Beneden); tache de Wagner, nucleole plasmatique, n. 
mixte, n. nucleinien, nucleole-noyau (Carnoy); nucleole adventif 
(Roule); corps nucleolaire, nucleolite (A. Schneider); nucleole 
primitif et secondaire (Carnoy and Lebrun); corpuscule ger- 
minatif (Van Beneden). 

Italian writers. — Macchia germinativa, macchia germinativa 
principale, m. g. laterale, m. g. accessoria. 

Synonyms of the nucleolinns. — Nucleololus, Nucleollolus 
(Frenzel); Schron’scher Korn, Valentinian vesicle, entostho- 
blast (Agassiz) ; Centrosoma (Lavdowsky); nucleolo-nucleus, 
endonucleolus (Macfarlane); Nucleolinus, Keimpunkt, punctum 
germinativum (Haeckel). 

III. OBSERVATIONS. 

A. Methods of Study. 

The following observations have been made upon material 
collected, fixed, stained, and sectioned by myself, with the 
exception of the preparations of the ova of Rodalia y which 
were kindly loaned to me by Dr. E. G. Conklin. In no case 
were observations made upon the living tissue ; however, but 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


401 


little could be gained from a study of the living cells, in regard 
to the minute structures with which we are chiefly engaged. 
With only few exceptions (Rodalia and the two gregarines 
examined) no cells were studied which had not been preserved 
with at least three fixing reagents, and in some cases at least 
half a dozen different fixatives were used. The preserving 
reagents employed were the following: saturated solutions of 
corrosive sublimate in distilled water (this being the only fluid 
used hot), sat. sol. of the same in 50 jo or 35 jo alcohol, Flem¬ 
ming’s stronger fluid (chromo-aceto-osmic acid), Hermann’s 
fluid (platinum chloride, acetic acid, osmic acid), sat. sol. of 
picric acid in 50 jo alcohol, Perenyi’s fluid (chromo-nitric acid), 
2 jo aqueous sol. of chromic acid, absolute alcohol, picro-nitro- 
osmic acid. Those reagents which gave the best general results 
were the fluids of Flemming and Hermann, and the alcoholic 
solution of corrosive sublimate ; though the particular reagent 
demanded depends both upon the object of study, as well as 
upon the method of staining which is to follow. It is hardly 
necessary to state that a structure found after the use of a 
given fluid, but not apparent on material treated in a different 
manner, was either regarded as an artefact, or doubts were 
expressed as to its naturalness ; that is, only when a structure 
was found to present itself to the eye in more or less the same 
manner, after various methods of preservation had been 
employed, have I regarded it as a natural appearance and not 
as a result of the fixatives used. Thin serial sections were cut 
of objects imbedded in paraffin, in the usual way. All staining 
done was upon the sections on the slide, and the stains employed 
were as follows : Ehrlich’s or Delafield’s haematoxylin followed 
by eosin (sat. sol. in distilled water), nigrosine (a sat. sol. in water 
diluted by six vols. water), sat. sol. of acid fuchsine in 50^ alco¬ 
hol, the triple stain of Ehrlich-Biondi-Heidenhain (as prepared 
by Griibler, Leipzig), Flemming’s triple stain (safranin, gentian 
violet, and orange G.), Lyons blue (sat. sol. in 50^ alcohol), 
gentian violet (sat. aqueous sol.), methylen blue (sat. aq. sol.), 
brasilin (sat. sols, in water and in 35^ alcohol), Mayer’s acid 
carmine, cochineal (sat. sol. in 70ft alcohol); while Grenacher’s 
borax carmine and alum carmine, Heidenhain’s iron haematoxy- 


402 


MONTGOMERY. 


[Vol. XV. 


lin, indigo-borax carmine (Norris and Shakespere), and certain 
others were tried, but proved unsatisfactory. With the excep¬ 
tion of the three triple stains mentioned, the others were used 
in various combinations as double stains ; worthy of recommen¬ 
dation are (with especial regard to the differentiation of the nucle¬ 
olus) Delafield’s, or better, Ehrlich’s haematoxylin followed by 
eosin ; acid carmine followed by nigrosine ; methylen blue fol¬ 
lowed by brasilin. Other combinations were also used, but it is 
not necessary to mention these here, nor to speak of the dura¬ 
tion of the staining baths, since in the explanation of the figures 
these data are given for each case separately. 

For the study of the finer structural details, the T ^th homo¬ 
geneous immersion lens of Zeiss was used, in combination with 
oculars 2 and 4. I would emphasize the fact that the drawings 
from the preparations were made gradually, as I proceeded in 
the study of each particular cell, and were not postponed until 
the end of the particular investigation, so that almost all were 
made before I had arrived at any views upon the nature of the 
nucleolus ; and I have pursued this method in order to elimi¬ 
nate from the figures as much as possible of the subjective ele¬ 
ment. In other words, I have made as close copies as possible 
of the preparations, drawing every cell or structure present¬ 
ing some appearance with which I had not as yet become 
acquainted, or rather the significance of which I had not 
learned, and then from the figures so made I have endeavored to 
learn the nature of the phenomena there presented, at the 
same time recurring to the preparations themselves. This 
method of study is the one employed by many investigators, 
though it can scarcely be termed the one most in vogue. The 
colors of the original figures have on the whole been most excel¬ 
lently reproduced by the lithographs of Werner and Winter. 

B. Protozoa. 

1. Gregarine from Lineus gesserensis (O. F. Miill). 

(Plate 21, Figs. 1-19.) 

(Description of the animal. —The largest individuals are just 
visible to the naked eye, and are of a whitish color. No 
synzigia were observed among the thirty individuals exam- 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


ined. Form : elongate, slightly larger at one end than the 
other, the thinner end sometimes flattened, slightly curved or 
sickle-shaped ; the greatest diameter is found in the region of 
the nucleus, which is situated nearer to the larger than to the 
smaller end ; both ends of the animal are rounded. In one 
individual (Fig. 2) the surface of the body was slightly furrowed 
in a spiral direction. Nucleus large, with a very thick mem¬ 
brane, and seldom oval, usually irregular in outline. In a single 
case (Fig. 1) two nuclei were present in one gregarine (the 
youngest individual seen), the two nuclei were of unequal size, 
though each contained a single nucleolus. Kolliker (’49) has 
described a gregarine with two nuclei ; I am unacquainted with 
any other cases. Sporocysts were not observed ; but in one 
case the cytoplasm was quite densely filled with minute spher¬ 
ical and oval bodies, which stained lightly with eosin, and in 
each occurred a small granule (this staining with haematoxy- 
lin) ; in the same individual a normal nucleus was also present 
(Fig. 4). These small bodies cannot be other than spores, even 
though they occur in the endoplasm of a gregarine in which 
a nucleus occurred at the same time; this observation stands in 
no accord with what has thus far been described of the sporu- 
lation among gregarines, and I am thoroughly at a loss to 
explain the phenomenon. These gregarines occurred only in 
the posterior intestine of Linens , but were not present in all 
the individuals of this nemertean sectioned. The absence of 
synzigia, the transverse furrows of the body, and the oval-shaped 
spores would relegate this form to the neighborhood of the 
genus Gonospora of Schneider.) 

In the smallest nuclei found (the size of the nucleus stands 
in some degree in proportion to that of the animal) only one 
nucleolus was present (Figs. 3 and 5) ; in all the larger nuclei 
their number varied from two to four, though since four nucleoli 
were found in only two cases, two or three nucleoli may be 
regarded as the usual number in the larger individuals. As 
an inspection of Figs. 3-19 shows, the comparative size of 
the nucleoli within the same nucleus is very variable, and the 
nucleoli of one nucleus are always of unequal size. When only 
two nucleoli occur, one is about one-half or three-quarters the 


404 


MONTGOMERY. 


[Vol. XV. 


size of the other; but when three nucleoli are present, either 
(i) one is particularly large, and the other two small; or (2) two 
are large, and the third is much smaller than either ; or (3) all 
three are large, the smallest being about one-half the size of the 
largest. In the two cases of nuclei with four nucleoli apiece, in 
the one there were two larger and two smaller nucleoli, in the 
other one large and three small ones. 

The nucleoli vary from a spherical to an oval shape. In the 
smallest usually no vacuoles (; n . Vac .) are to be seen, but such 
vacuoles are always to be found in the larger nucleoli. In the 
largest there is usually a large excentric vacuole, while small 
ones may or may not be present in other portions of the nucle¬ 
olus. In nucleoli of medium size it is most usual to find a 
number of small vacuoles. These vacuoles have already been 
noticed in numerous other gregarines, but I would call especial 
attention to a remarkable polarity of the nucleolus with regard 
to their position. In all those nucleoli in which vacuoles 
occurred, with the exception of not more than five or six, the 
single large vacuole, or the group of smaller ones, was situated 
at that pole of the nucleolus nearest the nuclear membrane 
(Figs. 7-9, 16, 17-19). There are almost no exceptions to 
this phenomenon in the smaller nucleoli, those, namely, in which 
only a single small vacuole or a few small ones are present. 
Accordingly, it would seem to be the rule that the vacuoles 
first appear in that portion of the nucleolus which approaches 
nearest to the nuclear membrane. The number and size of 
these vacuoles increase with the size of the nucleolus ; or, as 
is more usually the case, as the nucleolus increases in size they 
gradually fuse together to form a single large vacuole, which 
may occupy the greater part of the nucleolus (Fig. 15). Thus 
the vacuoles first arise at one point in the nucleolus, so that 
here one can speak of a polarity of the nucleolus ; but as the 
vacuoles increase in number and commence to fuse together 
the fluid substance of them begins to diffuse more widely 
throughout the nucleolus, so that evidences of this primitive 
polarity gradually become obliterated. 

The ground substance of the nucleoli is very finely granular, 
and stains deeply red with eosin, and brownish red with the 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


Ehrlich-Biondi stain. The vacuoles are filled with a structure¬ 
less fluid, which stains but lightly. But in four nuclei, the 
sections of which were stained in aqueous solution of methylen 
blue followed by brasilin, a differential stain of the ground sub¬ 
stance was acquired : that pole of the nucleolus which con¬ 
tained vacuoles was stained a bluish green (methylen blue), the 
opposite pole, where no vacuoles could be seen, being of a light 
pinkish color (brasilin), the vacuoles themselves appearing as 
clear unstained spaces (Figs. 17-19). In one nucleus, in which 
two minute nucleoli were present, the one without, the other 
with, a single small vacuole, both nucleoli stained a bluish 
green throughout (Fig. 18). Further, in an unstained nucleus 
fixed with Flemming’s fluid a somewhat similar differentiation 
was visible in the two larger nucleoli (neither of which con¬ 
tained vacuoles), the pole of each nucleolus nearest the nuclear 
membrane being of a deeper color than the opposite pole (Fig. 

11). This differentiation produced by staining would show that 
the ground substance of the smallest nucleoli is homogeneous, 
but that in the larger ones a chemical change takes place in it, 
whereby that portion of the substance opposite the pole where 
the vacuoles first appear differentiates itself chemically from 
that portion of the ground substance lying at the latter pole. 
Unfortunately I had too little material to carry further the 
study of this differentiation. 

In the nucleus is a faintly staining nuclear sap, in which 
irregular granules of various size are massed together espe¬ 
cially near the center of the nucleus ; they do not come into 
contact with the nucleoli, usually leaving a clear space around 
each of the nucleoli (Figs. 7, 8, 11, 14, 17-19). These do not 
stain with haematoxylin or with methylen green, but stain 
red with eosin and brownish red with the Ehrlich-Biondi mix¬ 
ture, in their staining differing little from the substance of the 
nucleoli. With the methylen-blue-brasilin stain mentioned 
above they stain pink, a little more deeply than does the inner 
pole of each of the larger nucleoli (Figs. 17-19). Whether they 
represent physiologically chromatin, or whether they are 
masses of (perhaps nutritive) substance taken into the nucleus 
from the cytoplasm, which might be chemically and genetically 


406 


MONTGOMERY,\ 


[Vol. XV. 


akin to part of the substance of the nucleoli, I am unable 
to decide. I am also unable to determine from the prepara¬ 
tions at hand whether the nucleoli themselves are partially 
composed of chromatin ; but the usual diagnostic stains for 
chromatin do not show the presence of this substance within 
the nucleus. 1 

To revert again to the polarity of the nucleoli. The fact 
that the vacuoles first arise in that portion of the nucleolus 
nearest the nuclear membrane would seem to prove that the 
substance of these vacuoles is extranuclear in origin, or else is 
secreted in the peripheral portion of the nucleus. But since it 
would be obscure how the peripheral portion of the nucleus 
should secrete a substance, and the central portion should not, 
I incline to the former explanation, namely, that the substance 
of the vacuoles is first produced in the cytoplasm, and then this 
substance penetrating through the nuclear membrane, it, or a 
part of it, arrives at that pole of the nucleolus nearest the 
nuclear membrane, and then is taken into the nucleolus at 
this pole. The size of the vacuoles stands in a more or less 
direct ratio to the size of the nucleolus itself; at the same time 
the ground substance of the nucleolus also increases in amount, 
though apparently not as rapidly as the amount of the vacuolar 
fluid. 

2. Gregarine from Carinella annulata . 

(Plate 21, Figs. 20-35.) 

{Description of the animal .—Monocystid gregarines occurring 
in the body cavity of this nemertean. No synzigia observed. 
Form : elongate, though not attenuate, the end in which the 
nucleus lies being broader and terminally more obtuse than the 
opposite end (Figs. 20 and 21). The longitudinal axis is never 
perfectly straight, and the cuticula shows no transverse fur¬ 
rows. The single nucleus is usually spherical or oval, rarely 
lobular in outline. In the entosarc of many individuals occur 
numerous minute, refractive granules. Neither cysts nor spores 
having been observed, I was unable to determine the genus of 

1 However, the chromatin here might exist in the state in which it is found in 
the growth period of ovocytes, namely, commingled with plastin. 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


this form. Only two individuals of Carinella were examined 
(both from Bergen, Norway); in the one all the gregarines were 
large, in the other of a smaller size.) 

The nucleoli are nearly always more numerous than in the 
preceding species of gregarine, the number varying from four to 
about twenty-six, in those stages found (Figs. 22-35). I n the 
larger nuclei they are usually more numerous than in the smaller 
ones, but exceptions to this rule are quite frequent. In the 
same nucleus some are nearly or quite spherical, others very 
irregularly lobular in outline. Their size within a given nucleus 
is also very variable, though as a rule they are unequal in their 
dimensions. In the larger nuclei the nucleoli are larger (or at 
least some of them are) than in the smaller nuclei. In a given 
nucleus there may be either (1) from two to four larger nucleoli 
and a number of smaller ones ; or (2) a single large nucleolus 
and several much smaller ones. In the smaller nuclei the 
nucleoli are more equal in size than in the larger ones. The 
largest nucleoli in a nucleus are as a rule of oval or spherical 
form, with regular contour (an exception is seen in Fig. 26) ; 
the irregularly lobular nucleoli (Figs. 23, 25, 27, 28, 33) are 
usually of medium or small size. There is no apparent regu¬ 
larity with regard to their distribution in the nucleus. None of 
the nucleoli appear to have limiting membranes. 

All these gregarines were fixed with alcoholic solution of 
corrosive sublimate. With the double stain, haematoxylin and 
eosin, the larger nucleoli were stained with a deep blackish 
red, the smaller ones either of the same color or a clearer red ; 
all became stained so intensely by this method that the vacu¬ 
oles in them were greatly obscured (Figs. 27 and 28). 

The Ehrlich-Biondi method produces a yellowish brown or 
reddish stain of the nucleoli, differences of stain being observ¬ 
able in the different nucleoli of the same nucleus (Figs. 26, 
31-35). This staining method brings out very clearly the 
vacuoles in the homogeneous (?) ground substance of the 
nucleolus ; the structureless substance of these vacuoles 
stains less intensely than the enveloping substance. Vacu¬ 
oles are absent in the smallest nucleoli, as well as in those of 
irregular form ; in the larger ones they are almost invariably 


408 


MONTGOMERY. 


[Vol. XV. 


present, though variable in size and number. They do not 
regularly arise at one particular part of the nucleolus, as we 
found to be the case in the preceding species. Further, there 
is rarely in this species a single large excentric vacuole ; but 
as the figures show, usually a number are present, either 
arranged in a circular row near the periphery, or in a row 
around a larger central vacuole, or grouped together at one 
point in the nucleolus. There can be no doubt that the larger 
vacuoles are produced by the fusion of smaller ones, since two 
or three smaller ones are frequently found in close contact 
with each other. 

The double stain, haematoxylin and alum carmine, gives 
different results from the preceding stains, in that by it not 
only the different nucleoli within a nucleus become colored 
differently, but also in some cases different stains of the 
different portions of the same nucleolus are attained (Figs. 
22-25). It is only the larger nucleoli, those with regular con¬ 
tours, which become differentially stained in this manner. In 
such a large nucleolus a portion of its substance stains a deep 
blue (haematoxylin), another portion or portions purplish or 
reddish (alum carmine) ; the part stained blue is usually central 
in position, and encircling it is a zone of red-stained substance. 
In one case (Fig. 22) the two opposite poles of the nucleolus 
were reddish, the intermediate part being a deep blue. The 
medium-sized, irregular nucleoli always stain blue throughout, 
the smaller ones usually red, but sometimes blue. This stain, 
accordingly, shows that in this gregarine some of the larger 
nucleoli are composed of two different substances similarly as 
we had found two substances in the preceding species, though 
there by using the methylen-blue-brasilin stain. 

With all three staining methods employed, a mass of irregular 
granules is present in each nucleus, which stain less intensely 
than the nucleoli. In the smallest nuclei (Figs. 22-25) these 
granules are more or less regularly distributed through the 
nucleus, but in the larger ones (Figs. 28, 31-35) they com¬ 
pose a dense mass around the nucleoli or around the largest 
nucleolus, while the peripheral portion of the nucleus remains 
nearly free of them. Delicate, faintly stained fibers transverse 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


this peripheral part of the nucleus, which may be radially dis¬ 
posed or else form a loose network. The size of the granules, 
their abundance and staining intensity vary in different nuclei 
of the same size, and there is no sharp distinction between the 
smallest nucleoli and the largest of these granules. In this 
species, as in the preceding, I was unable to detect any sub¬ 
stance which stained like chromatin. 

I have been unable to determine the origin and ultimate fate 
of these nucleoli, owing to lack of material ; but a few justifi¬ 
able conclusions may be drawn from the facts at hand. Thus 
the number and size of the nucleoli stand, as a rule, in a direct 
ratio to the size of the nucleus. Further, those irregularly lobular 
nucleoli described above probably represent amoeboid changes 
of the nucleolus, such as have been seen in life by previous 
investigators, though it is strange that these nucleoli differ from 
all others in consisting of a single substance and in containing 
no vacuoles. Lastly, the number and size of the vacuoles 
increase, as a rule, with the size of the nucleus. 

It is worthy of mention that usually there are a larger num¬ 
ber of very small nucleoli in the larger nuclei than there are 
in the smaller nuclei, although the largest nucleoli of the 
former are much larger than the largest nucleoli of the latter 
nuclei. We must conclude, then, that though the size of the 
nucleoli increases as a rule with that of the nucleus, new 
nucleoli are also being formed as the nucleus grows larger. 
Now some of these new small nucleoli found in the largest 
nuclei have undoubtedly been produced by division from some 
of the larger ones : thus I have frequently observed irregular 
(amoeboid) nucleoli with oval prolongations, or with small 
nucleoli closely apposed to their surfaces, and it probably is 
correct to conclude that such small nucleoli are in process of 
division from the larger ones (Figs. 23, 25, 27, 28, 33). 
Whether all the small nucleoli of the larger nuclei have 
had such a formation is difficult to determine, since in some 
of the largest nuclei most of the smallest nucleoli may be 
peripheral in position, close to the nuclear membrane, and 
far removed from the larger nucleoli, so that it might seem 
that the substance of these was extranuclcar in origin. The 


4io 


MONTGOMERY. 


[Vol. XV. 


mass of irregular granules within the nucleus appears to stand 
in some relation to the growth of the nucleoli, at least there is 
a relatively greater amount of this substance in the larger 
nuclei; it envelops the largest nucleoli and imbibes the same 
stains, though more faintly, with which the nucleoli become 
stained. Now as the gregarine grows, at the same time both 
nucleus and the total mass of nucleolar substance increase in 
size ; but the nucleus cannot grow without the addition of a 
substance or substances to it, which have been derived from 
without. Accordingly, I suppose that the substance of these 
granules has an extranuclear origin, a substance, i.e ., which, hav¬ 
ing penetrated the nucleus from the cytoplasm, undergoes a 
chemical change in the nucleus and there becomes precipitated 
in the form of granules, for no such substance occurs in granu¬ 
lar form in the cytoplasm. The growth of the nucleoli might 
then be explained on the assumption of the intussusception of 
this substance by the nucleoli. This explanation is offered 
merely as a hypothesis, since I cannot prove its correctness 
with the limited material at my disposal. Since no chromatin 
was demonstrable in these nuclei, it remains for future workers 
to show whether the chromatin is in these stages commingled 
with the nucleolar substance, or whether it is represented by 
one of the two substances of which some of the nucleoli are 
composed ; and if so, whether all, or whether only a certain 
number, of the nucleoli are thus partially constituted of 
chromatin. 1 


C. Metazoa. 

a. Egg Cells. 
i. Montagua fiilata (Verr.). 

(Plate 22, Figs. 57-63, 65-87.) 

In the germinal vesicles of this mollusc two kinds of nucleo¬ 
lar structures occur : the true nucleolus, which is of large size 
and almost invariably single ; and certain secondary structures, 

1 For observations of other authors on nucleoli in Gregarinida , cf. the reviews 
of Minchin (’ 93 ), Van Beneden (’ 69 ), Marshall (’ 92 ), Frenzel (’ 93 ), Koelliker 
(’ 49 ), A. Schneider (’ 75 , ’ 83 ), Wolters (’ 91 ), Camoy (’ 84 ). 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES . 411 

which appear at only a certain stage of the cell. The true 
nucleolus may be considered first, then these other structures, 
or “ pseudonucleoli.” 

There is always one true nucleolus to each nucleus, and in 
only two cases out of hundreds of ova examined have I seen 
two nucleoli (Figs. 57 and 61). The position of the nucleolus 
within the nucleus is in most cases excentric, seldom central, 
and never apposed to the nuclear membrane; it apparently lies 
free in the caryolymph, and is not supported by the chromatin 
threads. In the youngest, most immature germinal vesicles 
(I have not studied it in the ovogonia) it is apparently wholly 
homogeneous, dense, not noticeably refractive, and usually 
spherical (Figs. 57-61) ; sometimes, however, it shows an oval 
or more elongate form, and in the latter case its long axis 
usually coincides with that of the nucleus (Fig. 58) ; it is 
never irregular in outline. 

The nucleolus always colors differently from the chromatin, 
when treated with double stains, as follows : 


Stain. Nucleolus. Chromatin. 

Ehrlich-Biondi . . maroon . . . green. 

Haematoxylin, eosin . orange red . . blue. 

Acid carmine, nigrosine blue .... red. 
Haematoxylin, fuchsine purple . . . blue. 

Flemming’s stain . . yellow . . . violet. 


With the increase in size of the nucleus the nucleolus 
enlarges, and in such a way that the size of the latter usually 
preserves its proportion to that of the former ; but as the 
figures show, this proportion is quite frequently not preserved. 
What may be termed the first stage of this nucleolar growth 
consists merely in an increase in the amount of the homogene¬ 
ous substance, and between the largest homogeneous nucleoli 
(Fig. 65) and the smallest (Fig. 57) there is no difference 
except one of size. 

The second period of nucleolar growth is introduced when 
vacuoles commence to appear in the substance of the nucleolus 
(Fig. 62). Since my observations show that these nucleolar 
vacuoles are derived from small fluid globules which first appear 



412 


MONTGOMERY. 


[Vol. XV. 


in the nuclear sap, these globules may best be treated first. 
In the nuclear sap, at a certain stage in the growth period of 
the germinal vesicle, small globules of varying size occur ; 
there are usually one or two of them in a given nucleus, but 
sometimes they are quite numerous (Nut. Gl. in Figs. 62, 63, 
69-71, 73, 75, 81). When I first noticed these structures I 
conjectured that they might represent centrosomes such 
as have been found within nuclei at stages previous to mito¬ 
sis (by Brauer in the spermatocytes of Ascaris); but further 
investigation shows that they have no kind of relation to cen¬ 
trosomes, since they vary in number and size, and further they 
readily imbibe stains, which centrosomes do not. They have 
a close resemblance to the smallest yolk granules found in the 
cytoplasm in point of form, size, and manner of staining. How¬ 
ever, sometimes one or two of these bodies may be found in 
the nucleus when there is no evidence of yolk in the cytoplasm. 
Accordingly, they would seem to consist of a substance very 
similar to the young yolk at the time of its first formation. 
And since they may arise in the nucleus before yolk spherules 
appear in the cytoplasm they are probably not always taken up 
by the nucleus from the cytoplasm in the form of globules, but 
acquire this spherical form first in the nucleus. In other words, 
we may consider that the nucleus assimilates from the cyto¬ 
plasm a thin fluid, similar to, if not identical with, that from 
which the yolk spherules themselves are ultimately formed, and 
that in the nucleus this substance becomes deposited in the 
form of globules, perhaps after having undergone a chemical 
change within the nucleus. Further, this substance must be 
regarded as having a nutritive value, on account of its similarity 
to the substance of the yolk, which certainly is nutritive in 
function. In the more mature, larger germinal vesicles (Fig. 
78) large yolk globules are usually found, and are wholly simi¬ 
lar to those in the cytoplasm in these stages ; as can be easily 
determined, their position within the nucleus is not due to 
removal by the knife in sectioning, so that as the nucleus 
becomes larger it regularly takes up large yolk globules from 
the cytoplasm, and from these probably derives the greater 
part of the nourishment necessary for its rapid growth. We 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 413 

may conclude, then, that when the nucleus is comparatively 
small, and when no yolk or only small yolk globules are pres¬ 
ent in the cytoplasm, the nucleus derives a nutritive substance 
from the cytoplasm, which is closely similar to that composing 
the youngest yolk globules ; but when the nucleus has grown 
large, and the cytoplasm is packed with large yolk globules, it 
has the power to take up these larger globules also. 1 

To return, then, to the second stage of nucleolar differentia¬ 
tion. This stage does not commence when the nucleolus has 
attained a certain size, but may commence in some nucleoli 
earlier than in others; and again it is not marked by a particular 
stage of development of the yolk in the cytoplasm. The fluid 
vacuoles probably stand in a genetic relation to the small 
nutritive globules found in the nucleus, which have been 
just described. That is, these globules of the nucleus pene¬ 
trate into the nucleolus and then constitute the fluid vacuoles 
of the latter structure. I have reached this conclusion after 
observing that the vacuoles of the nucleolus and the small 
nutritive globules within the nucleus always stain in exactly 
the same way. This assumption is further strengthened by 
the fact that, when the nutritive globules lie in the nuclear 
sap at some distance from the nucleolus, they have invariably 
a spherical form ; but in those numerous cases where they may 
be seen apposed to the outer surface of the nucleolus they 
become flattened against the surface of the latter, as if the 
nucleolus were (figuratively speaking) a loadstone which 
attracts them to itself (Figs. 63, 69, 75). If this origin of 
the vacuoles of the nucleolus were not the true one it would 
be difficult to explain their mode of genesis, since there appears 
to be no other substance within the nucleolus from which they 
could be derived, and there is no reason for supposing that the 

1 The intensity in the staining of the yolk globules increases with their size, and 
the largest stain much more deeply than does the nucleolus. During all the 
earlier growth stages the nuclear membrane is retained, and it is seldom, and 
then only slightly, irregular in outline; therefore the yolk cannot be taken up by 
the mechanical aid of amoeboid processes of the nucleus, but its substance must 
osmotically penetrate the nuclear membrane. And as I mentioned above, it does 
not seem probable that the yolk globules retain their shape while penetrating this 
membrane, but diffuse through it in the form of an irregular fluid mass, and then 
in the nucleus this fluid becomes re-formed into globules. 


4 H 


MONTGOMERY. 


[Vol. XV. 


substance of these vacuoles is a differentiation of the nucleolar 
ground substance. We may assume, then, that this explana¬ 
tion of the genesis of the nucleolar vacuoles is the correct one, 
and now proceed to explain the changes in the nucleolus dur¬ 
ing the successive development of its vacuoles. If we take the 
size of the nucleolus as a general criterion (though it is not an 
infallible one, since there are considerable individual differences 
in different nucleoli (cf. Figs. 62, 65, 80)) of the stage of 
the nucleolus, the process of assimilation of the nutritive glob¬ 
ules from the nucleus by the nucleolus seems to be in general 
as follows : first, one or two globules are taken into the nucleo¬ 
lus, and later when others (apparently a varying number) are 
also taken up into it, we reach a stage when the nucleolus 
contains a number of fluid vacuoles (the assimilated nutritive 
globules) (Figs. 64 and 70). Then these vacuoles commence to 
fuse together (Figs. 63, 66, 72), finally by their fusion giving 
rise to one large vacuole, which fills about three-quarters of the 
space of the nucleolus, and always lies excentrically within the 
nucleolus (Figs. 68, 69, 73, 77, 79). The nucleolus has now 
attained its greatest dimension and is either perfectly spher¬ 
ical, or more usually ovoid in shape. Its large excentric vacuole 
is encircled by a peripheral layer of the primitive homogeneous 
ground substance of the nucleolus, which has undergone no 
structural or chemical change. This layer of ground sub¬ 
stance becomes necessarily thinner as the vacuole becomes 
larger, i.e., as the pressure from within becomes greater. But 
since the large vacuole lies peripherally, the peripheral sub¬ 
stance of the nucleolus remains thickened at that point opposite 
the vacuole, and this thickened portion of the nucleolar wall 
has most frequently the form of a concavo-convex lens (or on 
a cross-section, of a half moon), the concave side of which 
borders upon the vacuole. This thickened part, as the remain¬ 
ing portion of the peripheral layer of the nucleolus at this stage, 
is in every respect identical with the ground substance of the 
nucleolus in earlier stages, before vacuoles had made their 
appearance in it ; and the total amount of the substance of the 
peripheral layer seems to be equal to the amount of the homo¬ 
geneous substance of the nucleolus at the end of the preceding 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 415 

stage. Accordingly, in this second period of the nucleolar growth 
there appears to be no increase in the amount of the true 
nucleolar substance, but merely an increase in the amount of 
the vacuolar substance. The thickened portion of the periph¬ 
eral layer of the nucleolus is at first biconvex, but as the large 
vacuole grows larger the pressure of the latter causes it to 
gradually assume a concavo-convex form (Figs. 84-86). Thus 
the shape of the large vacuole is at first concavo-convex, and 
later spherical or oval. This thickened portion of the outer 
layer of the nucleolus is usually homogeneous in structure, as 
is the remainder of the true nucleolar substance which envelops 
the vacuole ; but sometimes small vacuoles may occur within 
it also (Fig. 71). 

Two poles may be distinguished in the nucleolus at this 
second stage of its differentiation : (1) the pole at which the 
large vacuole lies ; and (2) the pole at which the thickened mass 
of the true peripheral substance is situated. From the study 
of a large number of nuclei at this period I find that in about 
75j& of them the second of these poles is directed towards the 
nuclear membrane, the first pole towards the center of the 
nucleus ; at this stage, as in the preceding, the nucleolus lies 
usually excentrically within the nucleus. 

The later differentiation of the nucleolus consists, accord¬ 
ingly, in the accumulation in it of fluid vacuoles (their substance 
identical with that of the nutritive globules of the nucleus), 
but the true nucleolar substance undergoes no change whatever, 
as far as can be determined from differential staining. There 
is no chemical union of the vacuolar with the true nucleolar 
substance, but the fluid vacuoles simply push aside this sub¬ 
stance, so that, after these numerous smaller vacuoles have 
united to form a single large vacuole, the true nucleolar sub¬ 
stance remains unchanged as a peripheral layer around this 
vacuole. The substance of the vacuoles becomes colored with 
the same stains, though always more lightly, as does the true 
nucleolar substance, so that we find in this stage a more deeply 
staining envelope of substance around a less deeply stained 
portion. This difference of staining between these two parts 
of the nucleolus is best shown by employing haematoxylin 


416 


MONTGOMERY. 


[Vol. XV. 


and eosin (Figs. 68 and 69). With the Ehrlich-Biondi method 
this difference is not quite so clearly demonstrable. The latter 
stain is peculiar and differs from all other stains used by me 
for these cells, in that it very often gives to the smaller vacuoles 
of the nucleolus the appearance of black, refractive granules ; 
but a careful focusing of these supposed granules shows them 
without doubt to be vacuoles, their apparent solidarity being 
probably due to the refraction of light by the enveloping 
nucleolar substance. 

The chief result derived from the foregoing observations is 
that the nucleolus takes up some or all of those nutritive 
globules which lie in the caryolymph, and whose substance had 
been probably derived from the cytoplasm. Some of these glob¬ 
ules then become collected within the nucleolus, representing 
its fluid vacuoles ; and these globules, increasing in number at 
the same time, gradually fuse together and thus give rise to 
a single large excentric vacuole, which is enveloped by the 
unchanged true nucleolar substance. Since the substance of 
these small globules is probably nutritive in function, the 
nucleolus in thus collecting some or all of them would appear 
to act as a reservoir for nutritive substance, or as a reservoir 
for that portion of the nutritive substance accumulated in the 
nucleus, for which the nucleus may have no use. Of course 
it is not a priori impossible that these globules may represent 
waste products of a nutritive substance, so that the nucleolus 
might here fulfill the office of an excretory organ. But the 
function of these nucleoli can only be decided when the 
behavior of the nucleolus during the pole-body mitosis is 
known ; I had no ova showing pole-spindle formations. 

Finally, the true nucleolus appears not to be bounded by a 
special membrane ; after staining with acid carmine and nigro- 
sine the nuclear substance appears bluish green and a red 
membrane seems to envelop it (Fig. 80), but this appearance is 
probably due to the refraction of light, since nothing of the 
kind can be found after the use of other staining methods. 

We now come to speak of what I have called the “ pseudo- 
nucleoli/’ but merely in order to distinguish them from the 
true nucleolus, and without wishing to express by the use of 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 417 

this term any particular significance of these bodies. In eight 
individuals of Montagna which were sectioned, and which were 
of slightly different sizes, though the various growth stages of 
the ova were more or less the same in all, in only four were 
pseudonucleoli to be seen, and in only one of these four 
were they quite abundant, occurring in about 30 jo of the larger 
germinal vesicles. There are never more than from one to 
three in a nucleus. They are usually irregularly spherical and 
sometimes even angular in form ( Ps . n. in Figs. 72-77, 79). The 
largest attained about three-quarters the size of the true nucle¬ 
olus (of the same nucleus), though this size was attained by 
few, since they are, as a rule, but little larger than the nutritive 
globules which are observed in the caryolymph. Each pseudo¬ 
nucleolus consists of a denser, more deeply staining layer 
surrounding a less dense, more faintly staining core. The 
denser outer layer is homogeneous, somewhat refractive, and 
stains in the same manner as the ground substance of the true 
nucleolus. In smaller pseudonucleoli this outer portion appears 
on cross-section as a deeply staining ring, with regular out¬ 
lines, but in the larger ones small, irregular prominences may 
often be seen on its inner surface. The peripheral layer or 
ring, further, shows a double contour, but I am unable to deter¬ 
mine whether it is bounded by an outer membrane. It increases 
slightly in thickness with the growth of the pseudonucleolus, 
and in one case (Fig. 77) it was noticeably thickened at one pole, 
which gave to it somewhat the appearance of the tout ensemble 
of a true nucleolus. This peripheral layer surrounds a homo¬ 
geneous, non-refractive, probably fluid mass, which either stains 
not at all or else only faintly ; when it stains, it is either in the 
manner of the caryolymph or of the vacuoles of the true 
nucleolus. I have never noticed that the nutritive globules of 
the nuclear sap were apposed to these pseudonucleoli. What 
their origin is, and what their relation to the true nucleolus, I 
do not know. They are never found in contact with a true 
nucleolus and so are probably not buds from one. It is curious 
that they were frequent in the ova of only one mollusc, and in 
the same stages of the eggs of three other individuals were 
present in only a few cells, and in four other individuals were 


4 iS 


MONTGOMERY. 


[Vol. XV. 


present in none of the ova, though here the same stages of 
the ova were present as in the first individual. When they 
occur it is only in the larger germinal vesicles. They are 
apparently structures sui generis y and I have only the sugges¬ 
tion to offer, that they might be characteristic of a particular 
generation of egg cells, as their absence in the ova of some of 
the individuals of the mollusc would render probable (compare 
the observations of Hacker, ’ 93 a, where nucleolar differences 
were found in the ova of primiparous and multiparous individ¬ 
uals of Cyclops strenuus ). 

In Fig. 70 is a remarkable case depicted, namely, two small 
nuclei lying within a larger germinal vesicle, the former having 
apparently been assimilated by the latter. 

2 . Do to. 

(Plate 22 , Figs. 64 , 68 , 69 .) 

The nucleolar differentiation of these ova is essentially as in 
Montagna , so that no detailed description of the process need 
be given here. But in the five individuals of Doto which were 
sectioned, no traces of pseudonucleoli were seen, and the nutri¬ 
tive globules within the nuclear sap are usually smaller and 
much more numerous than in Montagna. The yolk globules 
also have different shapes in these two genera. 1 


3 . Amphiporus glutinosus (Verr.) 

(Plate 24, Figs. 140-158.) 

(For descriptions of the connective-tissue elements of the 
nemerteans, from which the genital products are derived, cf. 
my previous paper ’96.) 

In the nuclei of the connective elements, by a differentia¬ 
tion of which the ova are produced (without any intervening 

1 For the observations of other authors on molluscan germinal vesicles, cf. the 
reviews of the papers of Wagner (’35, ’39), Flemming (’74), O. Hertwig (’78b), 
Lonnberg (’92), Balbiani (’65b), Platner (’ 86 ), Leydig (’55a, ’50), Stauffacher 
(’93, ’97), Stepanoff (’65), Loven (’49), Mark (’81), List (’96), Blochmann (’82), 
Trinchese (’80), Heuscher (’93), Hubrecht (’81), Carnoy (’84, ’85), Wiren (’92), 
Fol (’89), Lacaze-Duthiers (’57), Quatrefages (’49). 


No. 2 .] COMPARATIVE CYTOLOGICAL STUDIES. 419 

mitosis), I could find no nucleoli ; but one or two small minute 
nucleoli might nevertheless be present within these nuclei, but 
escape detection, owing to their small size and to the compara¬ 
tively great amount of chromatin. These nuclei are usually 
elongated and irregular in form (Figs. 144 and 145, C. T. N.). 

The smallest germinal vesicles, which are recognizable as 
such by slightly larger dimensions and more regular, spherical 
shape, show likewise no recognizable nucleoli. 

In what may be termed the first nucleolar stage , the nuclei 
have grown still larger, and in them are to be seen from one to 
about twelve small nucleoli. These are all peripheral in posi¬ 
tion, being flattened against the inner surface of the nuclear 
membrane, which results in their not being spherical, but more 
or less flattened, lens-shaped, or hemispherical (Figs. 140 and 

141). 

Second nucleolar stage. — The peripheral nucleoli commence 
to wander towards the center of the nucleus, at the same time 
growing larger and increasing in number (Figs. 142-145, 152). 
This process goes on until a considerable number of quite large 
nucleoli are present, none of which are any longer in contact 
with the nuclear membrane. As a rule they are not evenly 
distributed throughout the nucleus, but groups of them occur 
at different points in the nucleus (Figs. 153, 146-150). This 
period of differentiation, then, consists in the grouping of most 
or all of the nucleoli at or near the center of the nucleus, 
accompanied by their increase in size. There is no ground for 
supposing that at this stage they fragment into smaller nucleoli; 
but very frequently groups of two or three nucleoli may be seen 
in close contact with one another, and these would represent 
states of fusion rather than of division, since they are found to 
be flattened at the point of contact, and not attenuated. Thus 
the increase in the size of the nucleoli would be due, in part at 
least, to fusion of contiguous ones. While some of the nucleoli 
have left the periphery of the nucleus, others are at the same 
time forming there, which in their turn eventually reach the 
center, so that a continual process of formation of nucleoli, and 
wandering of those already formed towards the center, takes 
place at this stage. 


420 


MONTGOMERY. 


[Vol. XV. 


Third nucleolar stage. —The nucleoli increase in number, but 
gradually become smaller and wander towards the periphery 
of the nucleus (Figs. 154 and 155), until they all lie close to 
the inner surface of the nuclear membrane. In this stage 
they attain their maximum staining intensity, as is well seen 
after the use of Heidenhain’s iron haematoxylin, by which 
they become colored a greenish blue (Fig. 157), while in the 
previous stages they are brownish yellow, unstained by the 
haematoxylin. 

Fourth nucleolar stage. — Vacuoles of varying size arise in the 
nucleoli, and become somewhat irregular (instead of spherical) 
in outline (Figs. 156 and 158). In numerous nuclei it may 
be noticed that all the nucleoli lie close to the nuclear mem¬ 
brane, except a single one, which is placed nearer the center 
and differs from the others in not staining with haematoxylin, 
though it usually contains vacuoles ; it may be a nucleolus 
which has not developed as fast as the others have (Fig. 
156). 

All nucleoli in the third and fourth stages are very uniform 
in size, and smaller and much more numerous than in the 
second ; since there are no facts which permit us to conclude 
that new nucleoli are being formed in the last two stages we 
must consider that in them a division of the nucleoli must take 
place, and this would explain their increase in number and 
concomitant decrease in size. The fourth stage would seem to 
be characterized by the commencement of a degeneration of 
the nucleoli, if the presence of vacuoles and the irregularity 
of form may be taken as a criterion of degeneration. Neither 
in this species nor in the other nemerteans examined have I 
seen stages showing the formation of the pole spindle, so that 
I cannot describe the ultimate fate of the nucleoli. But the 
observations of those who have studied these divisions seem 
to show that they all disappear before the pole spindles are pro¬ 
duced ; and accordingly the phenomena characteristic of our 
fourth nucleolar period might represent the commencement of 
these degenerative processes. 

The method of formation of the yolk may next be considered, 
since the yolk stands in a certain relation to the genesis of the 


No. 2 .] COMPARATIVE CYTOLOGICAL STUDIES . 421 

nucleoli. The cytoplasm, when the yolk first arises in it, 
stains with haematoxylin (with the double stain of this and 
eosin); this blue stain of the cytoplasm I have noticed to be 
characteristic for the cytoplasm of many immature ova, while 
the cytoplasm of somatic cells usually stains with eosin. The 
yolk first appears in the form of large yolk balls (Figs. 144 and 
145, Yk. Bl.)y as they may be termed; the number of these 
balls varies in cells of the same size, as well as in those of 
different dimensions, and they appear to be produced succes¬ 
sively in a cell, until at the end of the third nucleolar stage they 
all have disappeared, having given place to the mature yolk 
spherules. They arise in the cytoplasm at no fixed point, 
though usually at some distance from the nucleus ; it is hardly 
necessary to state that they stand in no genetic relation to the 
nucleus, either in this or in the other nemerteans studied. 
The yolk balls are at first dense and homogeneous, and stain 
intensely with eosin ; the size that they may attain while still 
homogeneous is very variable. Subsequently they become 
vacuolated, even sometimes granular, and different portions of 
the same ball may stain differently, which shows that both a 
chemical and a physical change takes place in their substance. 
Finally, they fragment into unequal sized granules, which stain 
less deeply, and then these latter split up further, until the 
ultimate yolk spherules ( Yk. Gl.) are produced. In the largest 
ovarial eggs all the yolk balls have disappeared (they linger 
longest at the periphery of the cell), the cytoplasm being 
densely filled with the yolk spherules. In some cases yolk 
balls lie in the cavity of the gonad (Fig. 155), and these are 
probably derived from degenerated ova. 

The following facts show, I think, that the nucleoli stand in 
a genetic connection with the yolk substance. The nucleoli 
stain in the same way and have in other respects the same 
appearance as the smaller fragments of the yolk balls and as 
the mature yolk spherules (Figs. 144-146). Fragments of 
• yolk balls occur frequently in close contact with the outer sur¬ 
face of the nuclear membrane. Now since the nucleoli first 
appear in contact with the inner surface of this membrane, the 
conclusion is plausible that the nucleoli represent portions of 


422 


MONTGOMERY. 


[Vol. XV. 


a yolk substance, either of the yolk-ball fragments or a sub¬ 
stance equivalent to that out of which the latter are differen¬ 
tiated, and this substance, then penetrating osmotically the 
nuclear membrane, becomes deposited or precipitated in the 
nucleus in the form of spherical globules, which are the nucleoli. 
From this yolk substance taken into the nucleus the chroma¬ 
tin, linin, and nuclear sap might derive the nourishment neces¬ 
sary for their growth, and those nucleoli which remain through 
the fourth nucleolar stage might represent either a reserve supply 
of this nourishment, or chemically changed portions of it, from 
which all nutritive substances have been extracted ; the latter 
view would seem substantiated by the fact that the nucleoli 
stain somewhat differently in the third and fourth stages. 

The nuclear membrane is present during all these stages. 
The nucleus is always regular in outline, usually oval, except 
during the third stage, when it may become slightly irregular, 
though it never becomes noticeably lobose or amoeboid. 

In the first nucleolar stage (Figs. 140 and 141) the chromatin 
appears as a network of delicate fibers, which stain with haema- 
toxylin. Towards the end of the second stage (Figs. 146-150) 
it assumes the form of irregular masses, and the fibers become 
less numerous. In the largest ovarial nuclei (Figs. 154 and 
157) it is finely distributed throughout the nucleus in the form 
of minute microsomes ; traces of fibers may be found only at 
the periphery of the nucleus, though I have not determined 
whether these are fibers now for the first time forming, as is 
the case in the other nemerteans. The nucleoli are never 
suspended by the chromatin fibers. 

This species is characterized by the formation of a mem¬ 
branous structure in the cytoplasm, during the second and 
third nucleolar stages, which is present in none of the other 
nemerteans. This is a membrane within the cytoplasm, 
separated from the nucleus, as well as from the cell membrane 
by cytoplasm; it lies close to the nucleus (Figs. 146 and 155, 
Iv. Mb.). It is thicker than the nuclear membrane, though 
not so dense, and differs in no wise structurally from the 
cytoplasm, except in its greater density, the cytoplasmic gran¬ 
ules in it lying closer together (these granules appear to be 


No. 2 .] COMPARATIVE CYTOLOGICAL STUDIES. 


the nodal points of a “ Wabenwerk ” in the sense of Biitschli). 
This intracellular membrane is not open at any point, and 
a longitudinal section of it shows it to be not spherical but 
oval in outline, the apices of the oval being furthest removed 
from the nucleus. It is present only in the second stage of the 
nucleolus, and between it and the nucleus no yolk balls occur. 
I have never seen such a structure in any other egg cells except 
in the ova of Gryllus abbreviatus ; a similar structure was found 
by van Bambeke (’ 83 , eggs of Leuciscus , Lota), Shafer (’ 80 , egg 
of Lepus ), and Gerould (’ 96 , Caudina egg). 

4 . Tetrastemma catenulatum (Verr.) Montg. 

(Plate 23 , Figs. 103 - 133 ; Plate 24 , Figs. 137-139) 

The formation of the yolk may be spoken of first, then the 
nucleoli proper, and afterwards certain large nuclear structures 
which may or may not represent nucleoli of another kind. 

The yolk first appears in the form of one or two yolk balls 
( Yk. Bl., Figs. 107, 108, 112, 114-116) in the cytoplasm; 
the larger ones are regularly oval as a rule, and the smaller 
ones spherical. A number of these yolk balls are produced 
successively in each cell, and by their fragmentation the ulti¬ 
mate yoke spherules ( Yk. Gl.) are evolved. Each such ball is at 
first smaller than the nucleus of the cell in which it occurs, but 
gradually increases in size, though the maximum size which it 
may attain is not a fixed quantity, but is quite variable. As it 
increases in size it also gradually becomes more deeply stained, 
attaining its most intense staining when it has attained the 
limit of size. The substance of these balls is dense, finely 
granular, not brittle, somewhat refractive ; in the youngest 
stages of their formation they often appear nearly homogeneous. 
About the time a ball has reached its maximum size it com¬ 
mences to change both structurally and chemically, vacuoles 
appear in it, it begins to stain less intensely, and becomes 
irregular in outline. Thus it becomes either coarsely granular, 
or else unstaining vacuoles appear scattered through it, and 
with eosin stains no longer a deep red, but a light red or even 
yellowish. Next it breaks into a number of pieces, whereby 


424 


MONTGOMERY. 


[Vol. XV. 


the primitive yolk ball may break either into two fragments 
(which are usually unequal in dimensions), each of which then 
fragments further, or it breaks at once into a considerable 
number of larger granules. The final stage in this process of 
division shows the daughter yoke balls fragmenting to form 
the ultimate yolk spherules (Fig. 118) ; the latter stain an 
orange red with eosin, are homogeneous in appearance, and 
usually oval or spherical in form, seldom irregular. Two main 
stages may accordingly be distinguished in the formation of the 
yolk : (i) the formation of a large, regularly shaped yolk ball ; 
and (2) the successive fragmentation of this ball, accompanied 
by a gradually lessening affinity for stains, resulting in the 
evolution of the mature, small yolk spherules, the cytoplasm 
of the ripe egg being thickly filled with the latter. It is usu¬ 
ally the case that the yolk ball attains its greatest size at the 
end of the first stage. In cells of medium size all the various 
stages of yolk formation may be found, which shows that the 
yolk balls are being successively produced and are successively 
fragmenting ; quite a number of these balls need to be pro¬ 
duced in order to furnish the large quantity of yolk globules of 
the mature egg. The time when the yolk balls first appear, 
the size they reach, and the manner in which they segment, 
seem to vary much in individual cells. 

I have not been able to determine the manner of the first 
differentiation of the yolk substance in the cytoplasm. Two 
possible explanations suggest themselves : (1) either a certain 
portion or constituent of the cytoplasm changes into yolk sub¬ 
stance ; or (2) the yolk balls may represent a nutritive 
substance accumulated in the cytoplasm, which may have been 
derived from the blood or from some neighboring tissue, if not 
directly from the posterior intestine. But it is without doubt 
that this substance is not of nuclear origin, for the yolk balls at 
their first appearance are not in contact with the nucleus, but 
usually at some distance from it ; and also during the earlier 
stages of the yolk formation the nucleus is irregular in outline, 
with short, blunt processes, which would show that it is tak¬ 
ing up substances from the cytoplasm, rather than excreting 
substances. 


No. 2 .] COMPARATIVE CYTOLOGICAL STUDIES. 425 


The cycle of the formation of the nucleoli may here also be 
divided into three stages, which do not quite correspond to the 
four of Amphipoms gelatinosus . 

First nucleolar stage. — In the smallest germinal vesicles 
found one or two relatively very large nucleoli were present, 
one of them often in the center of the nucleus, the other 
more excentric or even against the nuclear membrane (Figs. 
103, 114, 115). The nucleoli in these smallest nuclei are 
as large or nearly as large as in any of the following stages. 
In germinal vesicles of slightly greater dimensions three or four 
nucleoli may be present, and some of these may have increased 
a little in size ; the amount of nucleolar substance at this stage 
is often so great as to occupy a fifth of the nucleus. They 
now increase in number, until at the close of this period we 
find a considerable number of mostly large nucleoli quite evenly 
distributed through the nucleus (Figs. 104-106, 109, no, 116), 
but often they are at one of its poles more numerous than at 
other points. This stage would seem to correspond to the first 
and second of Amphiporus glutinosus. 

Second nucleolar stage . — The nucleoli continue to increase 
in number but now decrease in size and commence to pass to 
the periphery of the nucleus, until at the end of this period 
they all lie close to the nuclear membrane, are regular in out¬ 
line, and adequal in size (Figs. 107, 119, 122, 124-126, 130, 131). 
At the beginning of this stage numbers of nucleoli may be 
found arranged in chain-like rows, as is to be seen in Fig. hi. 
This would correspond to the third stage of Amphiporus. 

Third nucleolar stage. — Nearly all the nucleoli are close to 
the nuclear membrane, often flattened against it (Figs. 117, 120, 
127, 129, 137, 138). They show signs of degeneration; thus 
they stain less intensely, are irregular in outline, and have a 
vacuolar or granular structure. In the largest germinal vesicles 
their number has apparently decreased and small non-coherent 
masses of granules may be seen, which are probably degen¬ 
erated nucleoli. Sometimes a nucleus may be found in this 
stage in which almost all of the nucleoli contain each one 
large, excentric, lightly stained globule or vacuole (Fig. 117). 

Staining of the nucleoli. — The natural color would appear 


426 


MONTGOMERY. 


[Vol. XV. 


to be a light yellow. In a preparation stained with haematoxy- 
lin and eosin, though not very thoroughly colored with the 
latter stain, the large nucleoli of the first nucleolar stage were 
of a light-yellow color, apparently stained only slightly with the 
eosin; those of the end of the second stage were mostly stained 
red, and those of the third stage were stained red, except those 
which had broken into granules, these latter being stained very 
little. In another preparation, in which the eosin had acted 
for one or two minutes longer than in the preceding prepara¬ 
tion, the nucleoli in the first stage were stained orange, those of 
the second stage red, and those of the third stage very slightly 
or not at all stained. Accordingly, they stain more lightly at the 
commencement of the first and at the end of the third stage than 
during the second stage; these differences of stain are probably 
due to chemical differences in the nucleoli at different stages. 

The chief differences between the nucleoli of this species and 
those of Amphiporus glutinosus are as follows : in the former 
there is no stage which exactly corresponds to the first stage of 
the latter, where we found a number of small peripheral nucle¬ 
oli ; in T. catenulatum there are at first one or two large 
nucleoli which are not always peripheral in position. The 
nucleoli in the third stage of T. catenulatum are more irregular 
in form and dimensions and stain less intensely than those of 
the fourth stage of Amphiporus. But the most important differ¬ 
ence between the two species is to be found in the fact that 
in T. catenulatum new nucleoli continue to be produced even in 
the third stage. Thus there are at the periphery of the nucleus, 
between the larger degenerating nucleoli which had their origin 
during the first stage, also much smaller, newly formed nucleoli 
arising while the former are disappearing. Such younger 
nucleoli may be seen at the close of the third stage, when the 
nuclei are largest and chromatin filaments appear in them, 
arranged in contact with the chromatin threads or near to 
them (Figs. 127, 137, 138). These smallest nucleoli of the third 
stage always stain intensely red with eosin, while the much 
larger ones of the first and second stages stain more of an orange 
color with this stain. This difference of staining in these two 
kinds of nucleoli might be explained thus : 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


427 


As we had concluded for the preceding species, so also in the 
present and in the species of nemerteans yet to be described, 
the nucleoli are in all probability accumulations within the 
nucleus of a substance taken up from the cytoplasm, this sub¬ 
stance being related to that which constitutes the yolk balls. 
In the least mature germinal vesicles of T. catenulatum we 
found one or two very large, lightly staining nucleoli; these stain 
in the same way and show the same structure and degree of 
refraction as do the daughter yolk balls (Figs. 107 and 116). 
Further, I have noticed in the cytoplasm small yellowish spher¬ 
ules (yolk-ball fragments) which are in every way similar to the 
smaller nucleoli, and quite frequently I have observed one or two 
of them pressed so close against the outer surface of the nuclear 
membrane as to cause a depression of the latter (Figs. 112 and 
118). In other words, it would seem that the substance of 
some of the yolk-ball fragments is taken into the nucleus and 
in the latter is re-formed into nucleoli. As long as yolk balls 
or their fragments are found within the cytoplasm lightly 
stained nucleoli of approximately the same dimensions as these 
may be seen in the nucleus. I have never seen a pore in the 
nuclear membrane through which a yolk-ball fragment could 
penetrate, though this membrane sometimes appears to be 
thinner at the point of contact with a yolk-ball fragment than 
at other points in its circumference. But in the third stage, 
when all yolk balls and their fragments have disappeared and 
the whole cytoplasm is thickly filled with their derivatives, the 
mature yolk spherules, large, faintly staining nucleoli, are no 
longer present in the nucleus, but the smallest nucleoli present 
at this time resemble in form, size, and stain, the yolk globules. 
Therefore we must conclude that the young, small nucleoli 
which first appear about the end of the third nucleolar stage 
represent mature yolk spherules, or at least that the substance 
of the two is equivalent. While the nucleoli of the first gener¬ 
ation (formed in the first stage) are commencing to degenerate, 
new nucleoli of a second generation begin to arise in the 
nucleus, and the latter, which may serve as nourishment for 
the chromatin threads, differ from the former genetically, in that 
they are not assimilated portions of yolk-ball fragments, but 


428 


MONTGOMERY. 


[VOL. XV. 


assimilated yolk spherules. Thus, as we find in the cytoplasm 
first yolk balls, then their fragments, and finally the mature yolk 
spherules, so in the nucleus the first generation of nucleoli 
are assimilated yolk balls and their fragments, while the small 
ones of the second generation are derived from the only yolk 
elements then present in the cytoplasm, namely, yolk spherules. 
The nucleoli of the first generation also differ from those of 
the second, at the time of the first appearance of both, in 
their manner of staining ; so that they would seem to differ 
chemically from each other. 

Nuclear structures of problematical significance . — In only 
one out of the three individuals of this worm studied were the 
following remarkable structures to be observed, though the fixa¬ 
tion method of both of the other individuals was exactly the 
same. These bodies first appear in ova of the second nucleolar 
stage, but here show always the same typical structure, so that 
I can say nothing as to the manner of their first formation. In 
preparations stained with haematoxylin and eosin they are 
colored by the former stain a little more deeply than the 
nuclear chromosomes, so that they stand out sharply in the 
nuclear substance (N. Bd., Figs. 122-139). The smaller ones, 
i.e.y those of the younger germinal vesicles (Figs. 122-126), 
are finely granular, though whether they each consist of a mass 
of fine granules or of homogeneous ground substance in which 
granules are distributed, I cannot determine. In the larger 
nuclei they often appeared wholly homogeneous (Fig. 132). 
In shape they are usually nearly spherical, with a sharp outline, 
which may or may not represent a limiting membrane; the 
larger ones are often more irregular in form (Figs. 132, 133, 
139). In the smaller nuclei they are as a rule, but not always, 
smaller than in the larger ones ; in the smallest nuclei in which 
I have found them there is only one of these bodies to a 
nucleus ; while in the larger nuclei they are not only larger, 
but also there may be from one to four of them in each nucleus. 
In only one small nucleus were three of them present (Fig. 
128). In two cases, both larger nuclei, I found division stages 
of these bodies : in the one case (Fig. 131) the body was ovoid 
in outline, with a shallow constriction at right angles to its 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


429 


longitudinal axis, at about its middle ; in the other case 
(Fig. 129) the body was plainly biscuit-shaped, with a well- 
marked medial constriction : these would probably represent 
respectively successive stages of division. 

The various stages found would show the metamorphoses of 
these structures to be as follows : in the medium-sized nuclei, 
those in which they first appear, there is only one to a nucleus. 
This one increases in size up to a variable point, when it begins 
to divide, producing two daughter-bodies, which are not always 
of equal size. One or both of these bodies may now divide 
again, resulting in the formation of (respectively) three or four 
bodies. Since, however, the four bodies sometimes found in 
the larger nuclei are often quite unequal in size, we must 
assume : (1) either that the divisions have been very unequal, 
and each daughter-body had divided ; or (2) that after the first 
division, which may or may not have resulted in unequal 
daughter-bodies, only one of the latter divides further, and it 
divides once, and one of its products divides once. It is to be 
noted that the number, the size, and the time of the division 
of these bodies stand in no regular relation to the size of the 
nucleus. Thus in one small nucleus (Fig. 128) three were 
already present, so that here two divisions must have taken 
place; while in some much larger nuclei (Figs. 130 and 133) a 
single, much larger one was present, which showed no signs 
of division. In the larger nuclei these bodies are often quite 
irregular in form ; may this increasing irregularity portend 
an on-coming dissolution or other degeneration ? They were 
found, as remarked above, in the ova of only one of the three 
individuals of this species examined, though in all three indi¬ 
viduals the stages of egg development were very much alike; 
in the single individual in which they occurred they were not 
present in all the larger eggs. Their whole appearance and con¬ 
sistency show that they are not artefacts (the fixation was with 
hot aqueous corrosive sublimate), and they have no resem¬ 
blance to any parasitic organisms, as e.g.> Protozoa , with which 
I am acquainted. Nor can they be centrosomes nor true nucle¬ 
oli, and stand in no apparent relation to the nucleoli. In a 
single case I found two nucleoli enclosed by one of these bodies ; 


430 


MONTGOMERY. 


[VOL. XV. 


but in no other cases were these structures in contact with 
nucleoli. They are also never in contact with the nuclear 
membrane. Male pronuclei they cannot be, since the fecunda¬ 
tion takes place in later stages than those which I have had 
opportunity to observe. I must conclude, though with reserve, 
that they are either parasitic Protozoa , or, more probably per¬ 
haps, structures which characterize ova of a certain generation. 
(Compare my remarks on the “ pseudonucleoli ” of Montagna. 
The structure figured by Henneguy (’ 93 ), in the immature ger¬ 
minal vesicles of Sygnathus may have some connection with 
these bodies.) 

Chromatin. — The chromatin in the youngest germinal vesi¬ 
cles (Figs. 103-105, 112-114) is distributed throughout the 
nuclear sap in the form of minute microsomes. In the second 
and sometimes the first nucleolar stage such microsomes can 
often not be detected, but the whole nuclear substance, with 
the exception of the nucleoli, appears homogeneous and stains 
with eosin a yellowish red (Fig. 115). This peculiar coloration 
might be accounted for on the ground that in these stages there 
is a diffusion of nucleolar substance throughout the nucleus. 
Towards the conclusion of the second and the commencement 
of the third nucleolar stage, the minute chromatin microsomes 
again become evident (Figs. 118 and 130). At the end of the 
third stage a few chromatin threads begin to arise in the 
nucleus (Fig. 127), and these stain slightly with haematoxylin 
in the same manner as the microsomes do; they appear to 
arise separately and at different points in the nucleus, and are 
at first short, but gradually increase in length. As noted 
above, the small nucleoli of the second generation are often 
apposed to these threads, and sometimes lie in the meshes of 
them. 

Nucleus. — In the first and second nucleolar stages the 
nucleus has often short, lobular processes, which may be amoe¬ 
boid in life (Figs. 109, 112, 114, 116, 125) ; these changes in 
the form of the nucleus no doubt stand in a direct relation to the 
assimilation of yolk substance from the cytoplasm. Towards 
the end of the third stage the nucleus becomes regular in out¬ 
line, with no traces of amoeboid processes; at this stage also 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES . 


431 


the nuclear membrane has attained its greatest thickness. The 
thinness of this membrane in previous stages would allow the 
penetration of nutritive substances into the nucleus from 
the cytoplasm. The small nuclei from which the germinal vesi¬ 
cles are directly derived, without any intervening mitoses, are 
irregular in shape, and no nucleoli are to be seen in them 
(Figs. 108 and 112, C. T. N.). 


5. Tetrastem?na elegans (Verr.). 

(Plate 28, Figs. 282-299.) 

Having only two mature individuals of this worm for study, 
I am unable to give as thorough a description of the nuclear 
metamorphoses of the egg as was possible for the other nemer- 
teans ; one preparation was fixed with Hermann’s fluid, the 
other with aqueous solution of corrosive sublimate, but the 
latter had been too deeply stained (haematoxylin, eosin) to 
allow the study of certain details, as eg., the cytoplasmic 
changes leading to the formation of the yolk. Yolk balls were 
observed in only a few ova, and are much less numerous than 
in T. catenulatum ; it is possible that the development of the 
yolk in the present species may be as in Zygonemertes , that is, 
the mature yolk spherules may as a rule be directly formed 
without the interpolation of a yolk-ball stage. 

First nucleolar stage. — The youngest germinal vesicle, recog¬ 
nizable as such, showed a large nucleolus close to the nuclear 
membrane (Fig. 282) ; I have seen no smaller nuclei than this 
one, but would conclude by analogy from the facts in the other 
metanemerteans that also here all the nucleoli have an extra- 
nuclear origin. In slightly larger nuclei (Figs. 283-287) there 
are from one to three nucleoli, whose size varies considerably 
with regard to that of the nucleus, as well as to the size of one 
another. In such cases (Fig. 283) where only two nucleoli are 
present, one near the center of the nucleus, the other close to 
the nuclear membrane, the former is probably the older and 
has left the periphery for the center of the nucleus, while the 
other is younger and is still in process of formation. These 
first-formed nucleoli are usually rather large in proportion to 


43 2 


MONTGOMERY. 


[Vol. XV. 


the size of the nucleus, seldom small. It is the rule that in 
one, sometimes in all the nucleoli, a large unstaining globule 
is present, which has the appearance of a vacuole (Figs. 284- 
287, 298); no nucleolus has more than one such globule. Quite 
often there is only a single large vacuole-containing nucleolus 
in a nucleus ; or there may be from one to six nucleoli, only 
one of which contains a vacuole, and then the latter is usually 
the largest ; or again, there may be two or three large nucleoli, 
nearly equal in point of size, each of which contains a vacuole 
(of course numerous intermediate stages may be found). There 
is certainly a successive production of nucleoli, but it is diffi¬ 
cult to decide whether some of these after leaving the periphery 
of the nucleus fuse together, or whether some divide into smaller 
nucleoli. Now it seems probable that those nucleoli which are 
formed first are usually unequal in size, both in the same nucleus 
and in different nuclei, as a comparison of the figures shows. 
And though a gradual fusion of the nucleoli might play some 
part in the youngest germinal vesicles, nevertheless it would 
seem more probable that we have to do in these early stages 
with divisions of the nucleoli, especia^y since in the following 
stage they are much more numerous, as well as smaller. Fig. 
287, in which three apposed nucleoli are to be seen, may thus 
represent a division of a single nucleolus. It is not unlikely 
that the unstaining globule within a nucleolus might aid, if it 
is not the direct mechanical cause of, such division. This first 
nucleolar stage is then characterized by the successive forma¬ 
tion of a few comparatively large nucleoli at the periphery of 
the nucleus, and the migration of these towards the center ; 
the presence of large vacuoles within some of the nucleoli is 
also a criterion of this period. 

Second nucleolar stage .—We find a group of numerous 
nucleoli near the center of the nucleus, which are frequently 
more numerous than in our Fig. 292. At this stage they 
attain their smallest dimensions, and are approximately equal 
in size ; they are completely homogeneous and contain no 
vacuoles. The total number of the nucleoli is apparently 
greater at this stage than at any other. 

Third nucleolar stage. — This is characterized by an increase 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


433 


in the size of the nucleoli, a decrease in their number, and the 
gradual migration of them towards the periphery of the nucleus. 
At the beginning of this period (Figs. 290, 291, 293, 294), the 
nucleoli are quite evenly distributed throughout the nucleus; at 
its close they are mainly peripheral in position, near the nuclear 
membrane (Fig. 297). The increase in the size of the nucleoli 
is due, in some part at least, to the coalescence of every two or 
three neighboring ones, and such juxtapposed groups of two 
or three nucleoli may be often found (Fig. 294). None of the 
nucleoli contain vacuoles. 

Fourth nucleolar stage. — Now we find unstaining globules 
or vacuoles reappearing in the nucleoli, and there may be either 
a single large one to each nucleolus, or a number of smaller 
ones ; the large one is probably formed by the coalescence of 
smaller ones. Almost all the nucleoli are in contact with the 
nuclear membrane, often flattened against it (Fig. 299). They 
have become larger than in any preceding stage, and less 
numerous, but are now quite unequal in size. This stage may 
mark the commencing degeneration of the nucleoli, though I 
have observed no evidences of a commencing fragmentation. 

At the beginning of the first stage the nuclear sap never 
stains ; but at the end of this period, when the nucleoli have 
become more numerous, it stains very noticeably with eosin 
(Fig. 286), which would point to a solution of nucleolar sub¬ 
stance in the nuclear sap. 

6 . Zygonemertes virescens (Verr.) Montg. 

(Plate 27, Figs. 236-248.) 

Yolk. — In only two cases out of the numerous egg cells 
examined (three individuals of this worm were sectioned) have 
I seen yolk balls, so that the formation of yolk balls must be 
regarded as abnormal, if not pathological ; in this species the 
yolk arises as minute yolk spherules in the cytoplasm (Fig. 
246), without (except in the cases noted) a yolk-ball stage 
being passed through. These minute globules stain at first 
very faintly, and when they first appear are isolated from one 
another. There is no given point in the cytoplasm where they 


434 


MONTGOMERY. 


[Vol. XV. 


are first produced, but a varying number are formed simultane¬ 
ously and at different parts of the cell; it is usually, though 
not always, the case that they first arise at the periphery of the 
cell at some distance from the nucleus. The mature yolk 
globules are slightly larger than these and stain somewhat 
more intensely, which shows that they gradually become denser 
as they increase in size ; in the largest ova these spherules 
are so abundant that the true cytoplasm is quite obscured 
(Fig. 247). 

First nucleolar stage. — In the smallest nuclei found there is 
a peripheral group of several nucleoli lying close to the nuclear 
membrane, which are spherical in form (Figs. 236-238). 

Second nucleolar stage. —The nucleoli have increased in num¬ 
ber, and, departing from their original peripheral position, now 
occupy the center of the nucleus (Figs. 239 and 240). So small 
are they, and so densely grouped may they become, that at 
first sight one might be led to suppose that each group of 
numerous nucleoli was a single nucleolus. In those cases 
where the nucleus is oval or elongated in form, instead of 
spherical (the usual case), in the place of a single cluster two 
are commonly present, or else the single mass or cluster of 
nucleoli is elongate in shape, its outline being more or less par¬ 
allel to the contour of the nucleus. The nucleoli in this stage 
are always more numerous and usually also smaller than those 
of the previous period ; their increase in number might thus 
be brought about, in part at least, by divisions of the earlier 
nucleoli. 

Third nucleolar stage. —The nucleus now is much larger, 
and the nucleoli begin to wander apart towards the periphery 
of the nucleus (Figs. 241, 243, 246, 247). I have observed all 
stages between nuclei containing centrally grouped, small 
nucleoli and those in which they have come to lie close to the 
nuclear membrane. In this stage, as in the preceding one, 
the nucleoli are perfectly homogeneous without vacuoles, and 
spherical in form. In a few nuclei, however, they appear 
greatly vacuolated, but these cases are so rare that they must 
be considered abnormal. At the end of this period they attain 
their greatest dimensions, though they thereby become some- 


No. 2.] COMPARATIVE CVTOLOGICAL STUDIES. 435 

what unequal in size. In this stage, accordingly, they increase 
in size (perhaps by the fusion of contiguous ones (Fig. 242), 
and decrease in number, whereas in the preceding one the 
reverse process took place. 

Fourth nucleolar stage. — Almost all the nucleoli are flattened 
against the nuclear membrane (Figs. 245 and 248), and they com¬ 
mence to show a vacuolated structure; these apparent vacuoles, 
which are unstaining globules, when stained by the Ehrlich- 
Biondi method, whereby only the ground substance of the 
nucleolus is colored, appear as refractive granules (Fig. 248). 
At the conclusion of this period the nucleoli become irregular 
in shape, granular in appearance, stain less deeply, and each 
finally breaks up into a mass of granules. In this manner they 
decrease both in number and size. 

During the third and fourth stages, while the nucleoli are 
undergoing the metamorphoses described, a small number of 
newly formed ones appear in the nucleus, which are of later form¬ 
ation than the others (Figs. 245, 247, n. 2 ). These may serve 
as nourishment for the chromatic filaments, as in Tetrastemma 
catenulatum; but in the present species I have not observed 
any distribution of them along these filaments, and further 
they are numerically scarcer than in Tetrastemma. 

No yolk is present in the cytoplasm in the first and second 
nucleolar stages. This fact is easily proved by the use of the 
Ehrlich-Biondi stain, by which the cytoplasm is stained green, 
and the yolk substance, when present, a brownish maroon color. 
Yolk first appears in the third nucleolar stage, and at the com¬ 
mencement of the following stage the whole cytoplasm is nearly 
filled with it. Further, the nucleoli stain differently from the 
yolk globules by the use of the stain mentioned. These facts 
show that the origin of the nucleolar substance is not to be found 
in the yolk substance proper, but in a cytoplasmic substance 
from which the latter may later be evolved. That the sub¬ 
stance of the nucleoli is extranuclear in origin is shown by the 
fact that the nucleoli at their first appearance lie in contact 
with the nuclear membrane (Figs. 236-238), and only later do 
they take a central position. Though I have seen no nuclei 
smaller than those figured, which could without doubt be 


436 


MONTGOMERY. 


[Vol. XV. 


classed as germinal vesicles, yet it seems so probable that the 
substance out of which the nucleoli are formed is extranuclear, 
that I would conclude, a priori , that no nucleoli are present 
in stages of the germinal vesicle much earlier than those which 
have been here described. Those small nucleoli of a second 
generation, which are first produced in the third and fourth 
nucleolar stages, may represent yolk globules assimilated by 
the nucleus, since in these stages the cytoplasm is filled with 
such globules. 

On the other hand, the yolk cannot be considered as having 
its origin in nucleoli which have wandered out of the nucleus, 
since in none of these stages are nucleoli found in the cyto¬ 
plasm. And if such were the case, one certainly should be 
able to observe the large nucleoli of the third nucleolar stage 
in the cell substance, for it is at this period that the yolk first 
appears. I conclude that the yolk globules have their origin 
in some substance contained in the cytoplasm, and that the 
nucleolar substance also has its origin in some cytoplasmic 
substance. But whether the primitive nutritive substance of 
the yolk globules and that from which the nucleolar substance 
is derived are identical, is of course open to question ; how¬ 
ever, judging from the similarity in appearance, we might con¬ 
clude that the primitive cytoplasmic substance was the same 
in both cases, and especially if we consider, which seems plaus¬ 
ible, the nucleoli to represent the nutritive substance of the 
nucleus, as the yolk globules certainly represent that of the 
cell body. 

In the first nucleolar stage the nuclear membrane is usually 
very thin, but always perceptible ; in the later stages it becomes 
thicker. The nucleus is never noticeably irregular or amoe¬ 
boid in outline. Might this be explained by the absence of 
yolk balls in the cytoplasm ? 

In the second and at the beginning of the third nucleolar 
stages, the central mass of nucleoli is usually surrounded by a 
clear space, in which space few or no chromatin microsomes 
occur, though it may be transversed by a few achromatic fibers 
(Figs. 239 and 240). This space was found in most of the egg 
cells of this stage in the three individuals sectioned, though it 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


may have been produced by the action of the preserving fluid 
(hot aqueous solution of corrosive sublimate). 


7. Stichostem?na eilhardi (Montg.). 

(Plate 27, Figs. 213-235.) 

The yolk changes may first be delineated, then those of the 
nucleoli. In my paper on this fresh-water form (’95), I have 
described the ovogenesis to some extent, and here shall follow 
it more in detail. 

Yolk .— The yolk first appears in the cytoplasm in the form 
of small, more or less spherical masses (Fig. 213, Yk. Bl.), 
which at first stain like the cytoplasm ; but these youngest 
recognizable yolk balls consist of a substance in which the fine 
granules (or nodal points of an alveolar structure) are much 
more densely grouped than in the surrounding cytoplasm. 
Thus the young yolk ball may be distinguished from the 
cytoplasm proper by its greater density. A number of these 
yolk balls appear to arise simultaneously, though in these 
earliest, as well as in the later stages of yolk formation, a 
successive production and metamorphosis of yolk balls take 
place, since in all but the earliest stages of their development 
yolk balls occur in the cytoplasm in various stages of forma¬ 
tion. There is no rule as to the part in the cell at which these 
balls are destined to arise, for they may be found anywhere 
between the nucleus and the periphery of the cell; the fact 
that they first arise just as frequently at some distance from 
the nucleus as in its immediate neighborhood shows that they 
have no nuclear origin. An anabolic and a katabolic series of 
changes of each yolk ball can be distinguished, and these series 
of metamorphoses may be described in succession and termed 
respectively the prophasis and metaphasis of the yolk balls. 

Prophasis (Yk. Bl. in Figs. 217, 218, and the median ones of 
Fig. 215). — The progressive or anabolic changes of the yolk 
balls consist in (1) their absorbing protoplasmic stains with 
great intensity, so that they stand in marked contrast to the 
cytoplasm ; and (2) in their becoming quite homogeneous in 
structure, this homogeneity probably explainable by supposing 


43 8 


MONTGOMERY. 


[Vol. XV. 


that a dense condensation of the fine granules of which they 
are composed takes place. They continue to increase in size, 
and gradually stain deeper as they do so, until they attain about 
the dimensions given in Fig. 217 ; but I am unable to deter¬ 
mine whether they all reach exactly these dimensions before 
the metaphasic changes commence. At the conclusion of this 
period of their development they are large bodies, regularly 
spherical or oval in outline, and apparently without a limiting 
membrane ; especially characteristic is their homogeneity and 
their intense staining. 

Metaphasis. — These katabolic metamorphoses are intro¬ 
duced when a few unstaining globules arise in the substance 
of the yolk balls. These globules increase in number and size 
until the yolk ball assumes a vacuolated appearance (Figs. 215, 
217, 228). At the same time its ground substance loses its 
staining power and no longer stains homogeneously. At the 
commencement of these changes the yolk ball may even 
increase somewhat in size, since the substance of the globules 
is added to it. These changes continue until the yolk ball either 
breaks up into the mature yolk globules ( Yk . GY., Fig. 235), 
or first breaks into a varying number of larger pieces, and then 
each of the latter divides into yolk globules. The yolk globules 
are usually nearly spherical in shape, and though by no means 
equal in size are always larger than those of the other nemer- 
teans examined. 

During the prophasis each yolk ball is enveloped by a clear, 
structureless zone of cytoplasm ; but this surrounding zone is 
usually not noticeable around the larger yolk-ball fragments, 
and never around the mature yolk globules. 

As to the cause of the fragmentation of the yolk balls, I can 
find no sure explanation from the facts at hand. However, the 
appearance of the colorless fluid globules within their substance 
must have an important connection with these katabolic changes, 
since they characterize the commencement of this period of 
change. It would seem likely that these colorless globules 
represent a fluid constituent of the cytoplasm which has 
actively or passively been taken into the yolk ball, — perhaps 
from the clear cytoplasmic zone enveloping each yolk ball,— 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


since the yolk balls increase in size at the beginning of the 
metaphasis, though there appears to be no increase in their 
own ground substance. These unstaining globules are at first 
localized at different points in the yolk ball ; and it would seem 
probable that their substance later mixes itself with the ground 
substance of the yolk ball, since this supposition would account 
for the lessening intensity of the stain of the yolk ball during 
the metaphases. It would appear less probable that these 
globules are metabolic products of the true substance of the 
ball ; however, we have too few facts to enable us to deter¬ 
mine which of these is the correct view. 

Certain curious structures found in the cytoplasm of an 
immature worm fixed with Lang’s fluid (aqueous corrosive 
sublimate solution, NaCl, acetic acid) may be mentioned here. 
In the cytoplasm of a number of ova, in none of which any 
yolk was present, I found a few small, ring-shaped bodies, 
which stained with haematoxylin much more intensely than 
the surrounding cytoplasm ( Yk . Bl.f Fig. 233). Each con¬ 
sisted of a ring composed of a dense, homogeneous substance, 
the inner surface of the ring being smooth, but the outer irregu¬ 
larly jagged, this whole ring (as it appears on a section) enclos¬ 
ing an unstaining vacuole or globule. In reality these bodies 
are spheres, but my description applies to sections of them. 
These structures vary considerably in size, and sometimes are 
not spherical but oval, the larger ones usually staining more 
deeply than the smaller ones. I found them only in some of 
the ova of this one individual, and nothing of the kind was to 
be seen in the ova of about twenty other individuals sectioned, 
which had been variously preserved in picric, osmic, and chromic 
acids, in simple aqueous solution of corrosive sublimate, and in 
the fluids of Hermann and Flemming. Accordingly, they must 
be regarded as artefacts, produced by the action of the acetic 
acid, which I have long since found to be a very unreliable fixa¬ 
tive for the cytoplasmic elements of the nemerteans. It is 
most probable that they are young yolk balls, to which the 
acetic acid has given an abnormal appearance. Or might they 
represent multiple asters, such as have been recently described 
by Mead in Chaetopterus ? 


440 


MONTGOMERY. 


[Vol. XV. 


Germinal vesicles , nucleoli. — In this genus the earliest egg 
stages are more favorable for study than in the other metane- 
merteans. In the connective-tissue nuclei from which the ger¬ 
minal vesicles are directly derived (with no intervening cell 
generations) no nucleoli are present, though this conclusion was 
possible only after much careful observation. These small 
nuclei (Figs. 213, 217, 218, 220, 228, C. T. N.) are character¬ 
ized by a relatively thick membrane and by chromatin which is 
usually granular in distribution, but which may sometimes 
occur in the form of granular fibers. These chromatin masses 
might at. first sight be confounded with nucleoli, but their small 
size and irregular contours show that they are true chromatin 
granules. Further, when these nuclei are stained by the 
Ehrlich-Biondi method, these fibers and granules always stain 
with methylen green (chromatin reaction) and not a single one 
stains with fuchsine (which invariably stains any true nucleoli). 
Accordingly, what could not be finally proved for the other 
metanemerteans, though all observations pointed to its being 
the case there, could be definitely settled for Stichostemma> 
namely, that these connective-tissue cells contain no nucleoli ; 
in other words, nucleoli first arise in the definite germinal 
vesicles. 

Before proceeding to the description of the egg cells it may 
be noted that not all the undifferentiated connective-tissue 
cells within the gonad become germinal vesicles. I have previ¬ 
ously (’ 95 ) shown that the young gonad is a cell syncytium in 
which numerous nuclei are unevenly scattered through a mass of 
cytoplasm, but cell boundaries cannot be seen (Figs. 217 and 218). 
A few of these nuclei increase in size and eventually become 
germinal vesicles, and the latter reach maturity not simultane¬ 
ously but in succession, so that no gonad contains more than one 
large ovum at a given time. The numerous other nuclei which 
do not become thus differentiated degenerate, and their sub¬ 
stance is eventually absorbed by the gradually increasing mass of 
cytoplasm of one of the growing egg cells. These regressive 
processes are as follows (Fig. 218, C. T. N.) : the nuclei 
increase a little in size, but become much clearer in appearance, 
i.e., the relative amount of their chromatin appears to decrease ; 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 441 

next the cell membrane gradually disappears; then the chro¬ 
matin granules no longer become colored by any of the stains 
employed, but become refractive and yellowish. All the chro¬ 
matin granules do not lose their affinity for stains simulta¬ 
neously, but two or three of them may often remain stained as 
before, while the remaining granules of the same nucleus may 
have entirely lost their stain. At this period in the nuclear 
degeneration we find small masses of these unstaining, yellow¬ 
ish granules in the cytoplasm, each mass still preserving the 
form of a nucleus. Later these individual granules wander 
apart, or those of several nuclei may partially fuse together to 
produce a larger mass ; these larger masses of granules are 
always enveloped by a clear zone of cytoplasm, sometimes of 
considerable extent, so that they appear to be situated in 
vacuoles of the cytoplasm. The degeneration stages of these 
nuclei are most frequent in the cytoplasm, before yolk balls 
begin to arise in it ; as the latter appear, the remnants of the 
degenerated nuclei gradually vanish, so that when the cell is 
filled with the yolk balls all vestiges of these nuclei have 
vanished. We must suppose that they become assimilated by, 
or dissolved in, the cytoplasm. These formations, the katabolic 
changes of degenerating nuclei, can in no way be confounded 
with stages of yolk development, since the small size, yellowish 
color, and refrangibility of these granular masses serve to dis- 
- tinguish them sharply from any stage of the yolk balls, even 
though both are often found in the immediate vicinity of each 
other. 

The nuclei which are destined to become germinal vesicles 
increase in size to some extent before nucleoli appear in them; 
they now differ from the connective-tissue nuclei, apart from 
their greater dimensions, in having a relatively greater amount 
of chromatin and in being regularly spherical or oval in form. 
The first nucleoli to arise always lie in close contact with the 
inner surface of the nuclear membrane (Figs. 214, 216, 219, 
220, 224, 225). They usually appear in the form of a thin 
disc-shaped mass on the inner surface of the membrane, but 
there is considerable irregularity in the form of this mass, which 
may be angular or nearly spherical in outline. At the com- 


442 


MONTGOMERY. 


[Vol. XV. 


mencement of this first nucleolar stage the nucleolar substance 
appears at only one point in the periphery of the nucleus, and 
always in the shape of an irregular mass. 

Second nucleolar stage .—This period is characterized by 
the formation of other nucleolar masses at various points in 
the periphery of the nucleus, the successive detachment of all 
of these from their connection with the nuclear membrane, and 
their migration towards the center of the nucleus. The com¬ 
mencement of this process is to be seen in very young nuclei, 
where but a single peripheral nucleolar mass is present ; from 
the inner side of this mass small particles become divided off 
(Figs. 219, 224, 225), then each of these particles assumes a 
more or less spherical shape and wanders to the center of the 
nucleus ; this process continues until the whole mass of nucleo¬ 
lar substance has reached the center in the form of separate 
particles (Figs. 217, 218, 223, 227). The peripheral nucleolar 
mass usually stains less intensely than the portions which have 
already reached the center of the nucleus. While the first-formed 
peripheral nucleolar mass is thus gradually wandering to the 
center, other masses are successively forming at the periphery 
of the nucleus, and their detached portions successively passing 
to the center. When a considerable number of these nucleoli 
have reached the center of the nucleus they naturally come 
into mutual contact, and then a process of fusion sets in, which 
results in the coalescence of neighboring groups of nucleoli, so 
that a smaller number of larger ones are formed. Sometimes 
this fusion may proceed to such an extent that one single, 
enormous nucleolus results (Fig. 226), but usually several 
large nucleoli are the result, these being unequal in size. 
The irregularity both in the dimensions and the forms of the 
nucleoli is particularly characteristic for this stage ; thus the 
individual nucleoli often have elongated processes and angles, 
and this irregularity is frequently so excessive that the nucleoli 
within the nucleus appear like smears of ink upon a page (Figs. 
226, 227, 230). I think that this irregularity in form may be 
explained by the assumption that at this stage the substance 
of the nucleoli is viscid in its consistency, while in the follow¬ 
ing one, where the spherical form is the rule, its nature must 


No. 2 .] COMPARATIVE CYTOLOGICAL STUDIES. 


443 


be more freely fluid. Further, at this period we usually find 
vacuoles within some of the nucleoli of each germinal vesicle 
(Figs. 217, 218, 226, 229-231) ; sometimes no vacuoles are 
present in any of the nucleoli of a nucleus, but it is the rule that 
at least one of them, and that usually the largest, contains one 
or several vacuoles. Sometimes four or five of the nucleoli, 
which may be very unequal in size, may each have vacuoles. 
Occasionally a nucleolus contains only one vacuole, and in the 
latter there may be one or several small solid bodies, which 
stain like the ground substance of the nucleolus, and may be 
termed nucleololi; one of the latter may be fused with the inner 
surface of the nucleolar ground substance (Figs. 217, 218, 230, 
231). These nucleololi vary in number and size, and are 
absent in the greater number of the vacuoles ; so no particular 
significance should be attached to them, since they are probably 
nothing more than portions of the ground substance of the 
nucleolus which have become detached from the surrounding 
substance and have come to lie within the vacuole. During 
this period the nuclear membrane is thinner than at any other 
stage, and the nucleus is very noticeably amoeboid in form, the 
amoeboid processes being much more pronounced than in any of 
the other nemerteans examined; these processes in reality repre¬ 
sent changes in the form of the nucleus, and are not artefacts, 
since they are seen equally well after preservation in the most 
diverse fixing fluids (Figs. 226, 227, 230, 232, 233). The nu¬ 
clear membrane is always particularly thin around these nuclear 
processes, but, as far as I could make out, never becomes broken. 

Third nucleolar stage. — The large nucleoli which were 
present at the end of the preceding stage now commence to 
fragment into smaller nucleoli, which are more or less equal in 
size, and then the latter wander towards the periphery of the 
nucleus ; at the conclusion of this period, which must take 
place in a very short time, since I found only a few germinal 
vesicles exhibiting it, there are a large number of rather small 
nucleoli close to the nuclear membrane (Fig. 234). At this 
time the nucleoli attain their maximum staining intensity ; the 
nucleus usually shows no traces of an amoeboid form, and its 
membrane has increased in thickness. None of the nucleoli 


444 


MONTGOMERY. 


[Vol. XV. 


contain vacuoles ; and in every respect the nucleolar changes 
during this stage are the very reverse of the preceding. 

Fourth nucleolar stage. — This is characterized by the gradual 
degeneration and disappearance of the nuclei (Fig. 235). Small 
vacuoles arise in them, and these increase numerically, while at 
the same time the nucleolar substance stains less intensely. 
Fusion of neighboring nucleoli is very frequent at this time, or 
perhaps a little time before the nucleoli lose their staining 
power ; accordingly, in the largest germinal vesicles it is the 
rule to find a small number of large nucleoli. The nucleoli 
are not evenly distributed along the periphery of the nucleus, 
and are often flattened against the nuclear membrane. This 
nucleolar stage is found only in the largest ovarial eggs, where 
the nucleus is perfectly regular in outline, without amoeboid 
processes, and its membrane has attained its greatest thickness. 

Since this species is a protandric hermaphrodite, in which 
male and female sexual products ripen successively in each 
gonad, I found it at first difficult to determine whether a young 
nucleus in a given gonad corresponded to a male or to a female 
cell. But after comparing briefly the spermatogenesis of the 
other metanemerteans mentioned in this paper, and finding in 
them that no nucleus in any stage of spermatogenesis was larger 
than any of the smallest germinal vesicles here figured, I con¬ 
cluded that also in Stichostemma no male nuclei can attain the 
dimensions of even the smallest nuclei of our second nucleolar 
stage, and hence that all these nuclei were correctly concluded to 
be germinal vesicles, and not nuclei of spermato-genetic stages. 

We notice in the succession of the nucleolar stages described 
the rhythmic sequence in regard to (1) the position of the 
nucleoli, (2) their states of fusion and division, and (3) the 
absence and presence of vacuoles in them ; these successive 
changes may be expressed as follows : 

Nucleoli, 
a. 

t \ 

Position. Vacuoles. Fusion, division. 

peripheral . . . absent.fusion ? 

central . . . present . . . division, then fusion. 

peripheral . . . absent.division. 

peripheral . . . present . . . fusion, then division. 


Stage. 

First 

Second 

Third 

Fourth 





No, 2 .] COMPARATIVE CYTOLOGICAL STUDIES. 


445 


There is without doubt in this genus, as in the other meta- 
nemerteans, an extranuclear origin of the nucleolar substance. 
This is proved (i) by the absence of nucleoli in the nuclei 
from which the germinal vesicles are derived ; (2) by the 
nucleoli first appearing dose to the nuclear membrane. And 
since yolk globules do not arise in the cytoplasm until nearly 
the close of the second nucleolar period,when most of the nucleoli 
are near the center of the nucleus, to the yolk substance cannot 
be attributed a nucleolar derivation, and other reasons, such as 
the fact that the yolk balls usually appear at some distance 
from the nucleus, would contradict such an assumption. The 
nucleolar substance is apparently formed from an unstaining 
fluid constituent of the cytoplasm, which after it is taken into 
the nucleus undergoes a chemical change, since it stains there 
and is deposited in the form of nucleoli. In the second nucleolar 
stage, when the formation of nucleoli is at its height, the 
nuclear sap stains more deeply than at any other period (Figs. 
224-227, 233), so that it is probable that at this time nucleolar 
substance is finely distributed throughout the nuclear sap, as 
well as in the form of nucleoli. (This staining of the nuclear 
sap is especially well seen on material fixed with Flemming’s 
fluid and stained with alum carmine.) 

In the third and fourth nucleolar stages a few yolk globules 
are often found in a number of germinal vesicles (Figs. 234 and 
235, Yk. Gl .); these have probably been taken up by the nucleus 
from the cytoplasm. 

Chromatin. — In the nuclei of the first stage, the chromatin 
is always demonstrable in the form of coarse granules (Figs. 
214, 216, 219). In the beginning of the second it may usually 
be found in the form of a reticulation (Figs 218, 229, 233), but 
at the end of this stage it is not demonstrable (Fig. 227). In 
the third and fourth stages it reappears, but now in the form 
of fine microsomes (Fig. 235); and at the conclusion of the 
fourth stage short chromatic filaments begin to arise, similar 
to those described for Tetrastemma catcnulatum. 


446 


MONTGOMERY. 


[Vol. XV. 


8 . Lineus gesserensis (O. F. M.). 

(Plate 24, Figs. 159-177.) 

Yolk. — The yolk first arises in the cytoplasm in the form of 
irregular yolk balls, which are much smaller than in the other 
nemerteans examined (Yk. Bl., Figs. 159, 160, 177); these 
increase in number and size, the largest sometimes contain¬ 
ing vacuoles. In the largest ovarial ova seen (though I had 
only immature individuals of this species) yolk balls are no 
longer present, but in their place smaller yolk globules, which 
in all probability represent fragments of the earlier balls. The 
yolk usually makes its first appearance in a zone of the cyto¬ 
plasm, midway between the nucleus and the cell membrane, 
which is characterized from the rest by a less dense structure 
(Fig. 177). The extreme peripheral portion of the cytoplasm 
retains its density longest, as is also the case in the other 
species. The cytoplasm of the connective-tissue cells (Fig. 159), 
from which the egg cells take their origin, stains very faintly, 
while that of the young egg is dense and stains deeply. 

Nucleoli. — Only three worms out of eighteen sectioned con¬ 
tained ovogenetic stages, and since in these individuals only the 
earlier stages of this development were found, I am able to 
describe only the younger stages of nucleolar formation. The 
egg cell of this heteronemertean contains a single nucleolus ; 
apparent exceptions will be considered later. 

In the smallest nuclei (Fig. 159) of the cell syncytium of the 
gonads no nucleoli are to be seen ; we find nucleoli for the 
first time in cells whose nuclei are a little larger and whose 
cytoplasm commences to stain more intensely. These are the 
earliest stages of the ovocytes. 

Now in these youngest germinal vesicles (Figs. 159, 161, 
164, 166) the nucleolus is very frequently peripheral in position, 
close to the inner surface of the nuclear membrane ; while in 
the later stages (certain mitotic stages excluded) it is almost 
invariably never in contact with the nuclear membrane. Fur¬ 
ther, yolk balls first appear in the cytoplasm when the nucleus 
contains a nucleolus. These facts, being considered together 
with the fact that nucleoli are absent in the nuclei of the con- 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 447 

nective-tissue cells, lead to the conclusion that the nucleolus 
first appears in the young germinal vesicle, and more particu¬ 
larly, that the substance or che nucleolus is extranuclear in 
origin, and stands in a genetic relation to the substance of the 
young yolk balls. The substance of both is homogeneous and 
stains identically; by fixation in Hermann’s fluid, followed by the 
triple stain of Flemming, the nucleolus and the yolk balls stain 
a brownish yellow (Fig. 160); by fixation in corrosive sublimate 
and staining in haematoxylin and eosin both structures are 
colored a yellowish red (Fig. 177). Still more conclusive is 
the following observation : while the greater number of the 
yolk balls may lie at some distance from the nucleus, one or 
several are very frequently in close contact with the outer sur¬ 
face of the latter, and yolk balls may even be found which are 
halfway through the nuclear membrane, or which have com¬ 
pletely transversed it and lie within the nucleus (Fig. 160). 
Thus the nucleolus would seem to owe its origin to the sub¬ 
stance of yolk balls which have been taken into the nucleus. 
The very marked increase in the size of the nucleus and the 
nucleolus is probably caused by a continued process of yolk- 
ball assimilation on the part of the nucleus. This may be 
observed in numerous cases where small globules of yolk- 
ball substance lie within the nucleus, some at its periphery or 
close to the nuclear membrane, others flattened against the 
nucleolus (Figs. 160 and 177). By the use of the haematoxylin- 
eosin stain the nucleolar substance usually stains a little more 
intensely than the substance of the yolk balls (Fig. 177); this 
would show that this substance, after being taken up by the 
nucleus, undergoes a chemical change within the latter. Those 
yolk balls which are not assimilated by the nucleus remain in 
the cytoplasm and give rise to the yolk globules, as has been 
described. Thus the nucleolus probably has an extranuclear 
origin and represents a portion of the yolk-ball substance taken 
into the nucleus; its rapid increase in size is due to the addition 
to it of other similarly assimilated globules of substance. 

In the largest germinal vesicles seen (though these were 
not mature) the nucleolus is usually spherical in form, seldom 
oval, and homogeneous in structure, except that it sometimes 


448 


MONTGOMERY. 


[Vol. XV. 


contains a single large, unstaining globule, which appears as 
a vacuole (Figs. 162, 17 5-177) ; or there may be from one 
to three minute globules in it, which, when seen in their 
entirety, present the optical appearance (due perhaps to refrac¬ 
tion) of black granules, which might be mistaken for solid 
bodies. The nucleolus has no limiting membrane. The largest 
are relatively enormous and stain more intensely with eosin 
than the smaller ones. There is no clear zone in the nucleus 
around the nucleolus. 

In Linens the study of the metamorphoses of the nucleolus 
is complicated by the occurrence of nuclei in various mitotic 
stages. Karyokinetic figures were absent in the ovarial stages 
of the other nemerteans examined, so that in those species 
the connective-tissue nuclei and the egg nuclei both stand in 
the same cell generation, and the germinal vesicle may either 
be regarded as equivalent to an ovogonium or to a true ovocyte 
of the first order. In those species no cell generation separates 
the connective-tissue nucleus and the germinal vesicle, but the 
latter is merely evolved from the former by a gradual process of 
differentiation. But in Lineus the germinal vesicle is separated 
from the connective-tissue nucleus by at least one and prob¬ 
ably by two or three generations (if the differences in the size 
of the cells offer a sure criterion). Here, accordingly, the 
indifferent connective-tissue cell represents an ovogonium, and 
perhaps another generation of ovogonia may intervene before 
the germinal vesicle, the ovocyte of the first order, is produced. 
Of the two individuals on which these nuclear studies were 
made, I found mitotic stages in only one individual, while 
none were to be seen in the other individual, though here 
these nuclei had reached nearly the same degree of devel¬ 
opment. I have studied the mitosis merely with regard to the 
behavior of the nucleolus. The most abundant stages were 
those of the spirem and dispirem, asters and dyasters being 
much less frequent (Figs. 163, 166, 169, 170-172) ; the time 
duration of the latter stages may be less than that of the 
former. In by far the greater number of the spirem stages one 
nucleolus was present; it is probably present in each nucleus 
of this stage, but sometimes may escape observation by being 


1 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


449 


covered by the chromatic filament or by lying in a part of the 
nucleus outside of the plane of the section. In this stage, 
further, two nucleoli are never present; accordingly, in the 
spirem there is neither a disappearance nor a division of the 
nucleolus. In the dispirem stage each daughter-nucleus contains 
one nucleolus (Fig. 171), the two nucleoli being, however, often 
unequal in size. I found very few aster stages, and these were 
either so unfavorably placed for study or the chromosomes so 
densely entangled that I could not determine whether a nucleo¬ 
lus is present in this stage and whether a division of it takes 
place at this time. The facts determined are (1) that no divi¬ 
sion of the nucleolus occurs in the typical spirem stage, since 
here only one nucleolus is present ; and (2) that each nucleus 
of the daughter-spirem has one nucleolus. But I cannot show 
whether a division of the nucleolus occurs in the time between 
these two stages or whether the original nucleolus passes over 
into one of the daughter-nuclei, while in the other one a new 
nucleolus is produced. In these various mitotic stages the 
nucleolus usually lies at the periphery of the nucleus, and it is 
most frequently the case that it is not in contact with the 
chromatin filament ; it preserves its former shape and staining 
intensity, and apparently does not decrease in size during the 
mitosis. To be sure, in the karyokinetic stages under considera¬ 
tion it usually appears small in proportion to the size of the 
particular nucleus, but then it is usually the case in most 
mitoses, and probably so here, that before the disappearance 
of the nuclear membrane the volume of the nucleus greatly 
increases. 1 

Two nucleoli, never quite equal in size, are frequently found 
in certain small nuclei, which the distribution of the chromatin 
would show to be in a stage at the commencement of the 
prophasis of the mitosis or at the conclusion of the metaphasis 
(Figs. 163, 164-167, 170, 172). As the figures show, all these 
nuclei which contain two nucleoli are more or less of the same 
size. Nuclei which are a little smaller than these, as well as 
those which are larger, invariably contain a single nucleolus. 

1 The chromatin filament has considerable thickness and is apparently a con¬ 
tinuous thread ; it is looped around the inner surface of the nuclear membrane. 


MONTGOMERY. 


[Vol. XV. 


450 

It is probable that the two nucleoli of such nuclei have not 
arisen by division from a single nucleolus, but are nucleoli 
which have been developed at different points in the nucleus 
and which are destined to fuse together later and form a single 
one. This assumption was based upon the observation of 
nuclei where two nucleoli lie at opposite poles of a nucleus 
(Fig. 166) and each is apposed to the nuclear membrane, or 
where only one occupies such a peripheral position, the other 
being in the center of the nucleus (Fig. 164). In one figure 
(Fig. 165) we see a nucleus in which the two nucleoli lie near 
the center, close together, which might denote the beginning 
of such a fusion. On a little reflection this explanation of the 
presence of two nucleoli will appear quite allowable. In the 
more usual mode of development a larger nucleolus is formed 
at the periphery of the nucleus, wanders towards its center, and 
then much smaller masses of nucleolar substance are similarly 
formed and later fuse with the large nucleolus ; while in the 
cases under consideration two nucleoli of nearly equal size are 
produced, either simultaneously or in succession, and these 
afterwards fuse together. These two nucleoli of nearly equal 
size cannot be division products of a single primitive nucleolus, 
since two nucleoli are never found in the larger germinal 
vesicles. 

The nuclear sap of the smaller germinal vesicles does not 
stain at all ; in the larger ones (Figs. 168, 173—175, 177) it 
does, and the explanation for this staining may be given by the 
assumption that there is a dissolution of nucleolar substance 
throughout the whole nucleus, i.e., of that substance of the 
assimilated yolk balls which does not enter into the formation 
of the nucleoli. During the mitotic stages no constituents 
of the nucleus stain except the nucleolus and the chromatin 
filament, but these do not stain in the same manner. 

At first sight the heteronemertean Linens seems to differ 
markedly from all the metanemerteans here examined, in that 
it contains a single, enormous germinal spot. But in Linens , 
though a single large nucleolus is first formed, it nevertheless 
grows by the addition to it of much smaller nucleolar globules 
(Nut. 67 ., Figs. 168, 174, 177) which have the same method of 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 451 

formation and fuse with the former. Were these secondary 
nucleolar globules in Lineus as large as the first-formed 
nucleolus, and were they all to remain separate from one 
another, the nucleolar metamorphosis in this genus would 
correspond to that of the metanemerteans ; accordingly, the 
difference in the nucleolar production is not very important. 
(For the nucleolar relations in the other nemerteans, cf. my 
reviews of the papers of v. Kennel, Hubrecht, Coe and Burger. 1 ) 


9. Siphonophore (Rodalia .?). 

(Plate 26, Figs. 204-212.) 

(Dr. Conklin kindly loaned me the preparations on which his 
earlier studies were based (’ 91 ); these were preserved in alcohol 
and stained with haematoxylin.) 

There were no very young stages of the ovogenesis in this 
specimen ; I have studied the ova in the egg pouches and in 
the gonophores, each gonophore containing a single large ovum 
(as shown by Conklin and Brooks), while in the egg pouches a 
number of smaller ova may be present. 

A single large nucleolus is contained in each germinal 
vesicle. This is not only large in relation to the size of the 
nucleus, but is also absolutely probably one of the largest 
nucleoli ever described in animal cells (Fig. 212). It is always 
excentric in position, though seldom close to the nuclear mem¬ 
brane. In those younger stages where the nucleus is still near 
the center of the egg (Fig. 205, and the dorsal cell of Fig. 211) 

1 The only other observations of the yolk development in the nemerteans are 
those of Burger (’ 90 ) on Drepanophorus. Near the young germinal vesicle lies in 
the cytoplasm a homogeneous, deeply staining body, of smaller size than the 
nucleus, which Burger assumes may correspond to a yolk nucleus. This body 
disappears, “ und es sammeln sich namlich, dem Keimblaschen anliegend, in jenem 
[Plasmahiigel] kuglige oder langliche, tropfchenahnliche Gebilde an, erst sp'arlich 
ein einziges, zwei und mehrere, spater aber mit dem immer noch fortschreitenden 
Wachstum des Keimblaschens sich zahlreich vermehrend in grosster Menge. Sie 
sind durchaus homogen, von mattem Glanze und ausserst tinktionsfahig. . . . 
Erst nach der Entwicklung des Keimblaschens geht die des Deutoplasm as vor 
sich und zwar nun auf Kosten der glanzenden Dotterballen, welche aufgebraucht 
werden und so im reifen Ei verschwinden.” In the ripe egg the cytoplasm is 
granular and stains lightly. 


452 


MONTGOMERY. 


[Vol. XV. 


the nucleolus is usually nearer the center of the nucleus than 
in those more mature stages where the nucleus lies near the 
periphery of the cell. But in the more mature stages the 
nucleolus may lie at the animal pole, or the vegetal pole, or 
at one side of the nucleus, so that no coincidence between 
the position of the nucleolus and the age of the nucleus can be 
determined. Thus the nucleolus stands, eg., in no relation to 
the animal pole of the more mature nucleus, that pole where 
amoeboid processes are produced (Figs. 204 and 209). The 
ground substance of the nucleolus is dense and homogeneous, 
and stains quite deeply; the nucleoli of the smaller germinal 
vesicles stain, as a rule, less intensely. In the ground sub¬ 
stance of all the nucleoli more or less numerous fluid globules 
occur, which stain very faintly or not at all, and their presence 
gives a vacuolated appearance to the nucleolus ; those within 
the same nucleolus are of unequal size, and among them two 
or three usually occur which far exceed the others in size. 
Occasionally there is one large central vacuole (Fig. 206), but 
as a rule the larger ones are peripheral, and may produce prom¬ 
inences of the surface of the otherwise perfectly smooth and 
spherical nucleolus (Figs. 209 and 212). In one large vacuole 
(Fig. 212) a finely granular mass was found, though this may 
have been an artefact. Since in the smaller nucleoli these 
vacuoles are less numerous and smaller in size, it would seem 
probable that in stages antecedent to those found by me the 
nucleolus may be wholly devoid of such vacuoles. The nucleo¬ 
lus has no enveloping membrane, for what at first view appears 
to be such a structure careful study shows to be merely the 
result of refraction. 

In addition to the single large nucleolus described, there are in 
the most mature nuclei also from about one to five minute nuclei 
(Fig. 209). These vary somewhat in size, are perfectly spher¬ 
ical and homogeneous, without vacuoles, and stain more deeply 
than the larger one. Sometimes they are found in close con¬ 
tact with the nuclear filaments (cf. the nucleoli of the second 
generation in Tetrastemma catenulatum and the observations 
of Riickert (’ 92 ) on the germinal vesicles of Selachii). These 
probably have no genetic relation to the large nucleolus, since 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


they never lie in contact with the latter and are frequently 
situated at some distance from it. Were they buds from the 
large one, one would expect to find in them vacuoles such as 
occur in the large nucleolus, but they never contain vacuoles. 
In one nucleus (Fig. 207) I saw a disc-shaped mass apposed to 
the inner surface of the nuclear membrane, which stained more 
intensely than the chromatin. Such a peripheral mass may 
be regarded as a substance taken up from the cytoplasm by 
the nucleus, which, after passing through the nuclear mem¬ 
brane, undergoes a chemical change to such an extent that it 
stains with haematoxylin. The minute nucleoli may stand in 
a genetic connection with such a mass of substance, that is, be 
portions of a substance assimilated by the nucleus and after¬ 
wards scattered through the latter. They might serve as 
nourishment for the chromatin threads with which they are 
often in contact. 

In seven nuclei out of about one hundred or more examined the 
large nucleoli differed much from the ordinary type described 
above. In one egg pouch there was a smaller ovum apposed to 
the animal pole of a larger one (Fig. 211); a normal nucleolus 
was present in the nucleus of the smaller one. But in the larger 
ovum two nuclei were present, in close contact with one another, 
though separated by a membrane (coalesced nuclear membrane). 
It is in each of these latter nuclei that an abnormal nucleolus 
is present. Each of these nucleoli is finely granular, without 
enclosed vacuoles, and stains faintly with haematoxylin ; the 
one is regular in outline, but the other is jagged at one pole, 
and a ring-shaped portion of its substance stains more deeply 
than the remaining portion. In another ovum I also found two 
nuclei, in each of which was a nucleolus similar to those just 
described. In still another ovum two nuclei were found in 
contact with each other, the nucleolus of one of which was 
similar to those here described, but the nucleolus of the other 
nucleus was intermediate in structure between these and the 
ordinary type of nucleoli (Fig. 210). In only one case was 
such an abnormal nucleolus present within an ovum contained 
in a gonophore (Fig. 208); in the other six cases the abnormal 
nucleoli were in ova of egg pouches. 


454 


MONTGOMERY. 


[Vol. XV. 


Now what do these lightly staining, granular nucleoli repre¬ 
sent ? In all except the seven cases here mentioned the nucleo¬ 
lus was always of the deeply staining, vacuolar type, irrespective 
of its occurrence in ova of egg pouches and of gonophores. 
The abnormal nucleoli, with one exception, were found in the 
largest ova of the egg pouches. Types intermediate between 
the two are represented in Fig. 210. Conklin and Brook’s 
observations, which I can corroborate, show that a number of 
ova are produced in an egg pouch, but that only one of these 
passes into a gonophore, and there develops into the ripe ovum, 
while the others remain behind in the egg pouch and do not 
reach maturity, but degenerate. I would hold that the abnormal 
nucleoli described by me are degenerating nucleoli of degener¬ 
ating ova. All the facts seem to favor such an explanation. 

The cytoplasm of the youngest egg cells appears finely granu¬ 
lar (it may be an alveolar meshwork). In the largest it was 
coarsely vacuolar, especially near the center of the cell; I find 
no evidence of yolk. Conklin and Brooks evidently mistook 
the vacuoles of the cytoplasm for yolk globules. 

No chromatin threads were apparent in the smallest germinal 
vesicles (Figs. 204-206), but only a fine granulation in the 
nuclear sap ; chromatin threads make their appearance gradu¬ 
ally in the larger ova (Figs. 207, 209, 211) and stain more 
intensely as they increase in number and size. Each thread 
often has the form of a chain of transversely placed discs ; or 
sometimes it would seem to consist of a large number of short 
fibrils, placed at right angles to a common longitudinal axis, as 
is the structure of the chromosomes of the Selachian egg. 
These threads usually make their first appearance in the neigh¬ 
borhood of the nucleolus, from which they sometimes radiate 
outwards ; only in the largest nuclei are they more generally 
distributed throughout the nucleus. This fact might show a 
physiological relation between these two structures. But there 
is in all probability no genetic connection between the two ; 
rather, the chromatin threads are built up of the minute micro- 
somes found in the nuclear sap of the smaller ova. But the 
formation of the chromatin threads must be determined by the 
investigator who has more abundant material at his disposal, 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


and material which has been more advantageously fixed and 
stained. 1 


io. Poly dor a. 

(Plate 28, Figs. 249-281.) 

The egg cells of this form, as those of most Polychaeta , are 
derived from the peritoneal cells of the body cavity, the latter 
cells building pseudoepithelia around the intestine, as well as 
occurring free in the body cavity. Those in the pseudoepi¬ 
thelia (Fig. 249) are more or less flattened, disc-shaped, while 
the free cells (Figs. 250-254) are oval in shape, with more 
regular outlines. Their cytoplasm is not dense, and one or several 
large vacuoles are frequently found at the periphery of the cell; 
a delicate cell membrane is present. The cytoplasm of these 
sexually indifferent cells does not stain with haematoxylin. The 
nucleus is small, irregular in outline, and contains a few chromatin 
granules ; very frequently the greater part of this substance lies 
close to the nuclear membrane. I have never found more than 
one minute nucleolus, and this is almost always close to, or in 
actual contact with, the nuclear membrane (Figs. 251, 252, 254); 
in many nuclei I failed to find nucleoli, though in these cases they 
may have been obscured by the chromatin. I found one divi¬ 
sion stage of a nucleus (Fig. 249) ; there were two daughter- 
nuclei of the same size and form lying close together ; the 
nucleolus of each was somewhat elongate in form (in all others of 
these cells examined it is spherical), which might show that the 
nucleoli had been produced by the division of a single one in the 
mother-nucleus. In many of the smaller free peritoneal cells 
a peculiar body often occurs (N. P. Fig. 253). This is always 
smaller than the nucleus, more or less spherical, often homo¬ 
geneous in appearance, and it may stain either deeply red with 
eosin or faintly with haematoxylin, or in other cases it may not 
stain at all, but appear as a light yellowish, refractive mass. 
From the comparative study of a large number of cells contain¬ 
ing these bodies it may be determined that they are degener¬ 
ated nuclei or portions of nuclei. Thus in Fig. 250, which 

1 For other observations on nucleoli of Siphonophora , cf , besides the paper by 
Conklin and Brooks, the review of O. Hertwig (’ 78 b). 


456 


MONTGOMERY. 


[Vol. XV. 


probably represents the commencement of such a degeneration, 
there lies close to the nucleus what seems to be a much smaller 
nucleus, or a portion of one; and I have found all intermediate 
stages between such a body, which is granular and stains with 
haematoxylin, and the body reproduced in Fig. 253, which 
appears nearly homogeneous and stains with eosin. These 
bodies then seem to be degenerated or cast-off portions of 
nuclei. We might conclude also that the cells in which these 
structures are found, are themselves fated not to develop into 
egg cells, even if they are not degenerating; for no such bodies 
are to be seen in the cytoplasm of the true egg cells. 

These peritoneal cells have the morphological value of ovo- 
gonia. Those which are destined to become ova seem to 
become detached from the pseudoepithelial connection, but in 
such a way that they do not become detached singly, but 
portions, each of which is composed of a number of cells, 
become loosened from the epithelium. Thus the earliest ovo- 
genetic stages are to be found in strings of cells arranged 
radially around a common longitudinal axis, each such string of 
cells situated free in the body cavity (Fig. 270 represents a 
portion of such a string). At the one end of such a cellular 
string lie, densely grouped, the numerous mitoses of the 
ovogonic stages, while the remaining portion of the string is 
usually composed of young ova, sensu strictiori. I have never 
found mitoses in cells which lie singly in the body cavity. 

The first change noticeable in the ovogonium leading to the 
formation of the ovum consists in (1) the increase in the size 
of it and of its nucleus, and (2) in its cytoplasm gradually stain¬ 
ing with haematoxylin. This deep blue staining of the cyto¬ 
plasm, accompanied by its increasing density and the loss of the 
vacuoles in it, continues from now on until yolk granules begin 
to arise in it, when the cytoplasm commences to stain faintly 
with eosin and loses its dense structure. At the conclusion of 
the ovogonium rest stage the nucleolus has increased a little 
in size, accompanying the growth of the nucleus. 

The next stage is a mitosis. Whether there is more than 
one mitotic ,generation separating the ovogonium from the 
ovum I have not been able to determine; the slight differences 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


in the size of the mitoses hardly afford a satisfactory criterion 
for deciding this point (Figs. 255-261). All the typical stages 
of the prophase and metaphase are to be found, though only in 
the arrangement of the chromatin, for I have been unable to 
find either centrosomes or achromatic spindle. After careful 
study of a large number of these dividing nuclei I find the 
nucleolus to persist in the nucleus throughout the mitosis. 
Further, it appears to retain its original size throughout this 
process, without any diminution in volume. Thus the nucleolus 
seems to be retained without change in the spirem and aster 
stages of the prophasis. In the dyaster stage (Fig. 258) each 
pole of the nucleus usually contains a nucleolus, so that the 
nucleus contains two nucleoli ; and when the nuclear divi¬ 
sion is completed, i.e . 9 when in one and the same cell two 
nuclei occur in close contact with each other, in the aster as 
well as in the spirem of the metaphasis, each daughter-nucleus 
has its own nucleolus (Fig. 257). Now the ovogonium contains 
only one nucleolus, so that we must assume either (1) that a 
division of the nucleolus has taken place during the mitosis, or 
(2) that to one of the daughter-nuclei is allotted the whole 
original nucleolus, while in the other nucleus a new one is pro¬ 
duced. I have not seen any dividing nucleoli in these mitoses, 
their small size being a great obstacle to their study. But I 
should judge that such a division occurs, for these reasons : 
(1) the nucleus of one or of both the daughter-nuclei has 
sometimes a somewhat elongate form (Fig. 257); and (2) in 
later stages of the ovum proper I have found dividing nucleoli, 
and these cases would show that if such divisions take place in 
stages subsequent to the mitosis they might also occur during 
the mitosis. The two cases of division of the nucleolus found 
are here figured (Figs. 264 and 265), and in each of the elongate 
nuclei is a dumbbell-shaped nucleolus lying in the longitudinal 
axis of the nucleus ; in these figures the two halves of each 
nucleolus appear unequal in dimensions, but this is so because 
neither of these nucleoli happened to lie wholly in the plane of 
the section. I have found numerous other cases of elongated 
nuclei, each with an elongate nucleolus without any median 
constriction (Fig. 270). These facts would show that a division 


458 


MONTGOMERY. 


[Vol. XV. 


of the nucleolus may take place during the mitosis, and 
probably does so. 

After the completion of the mitosis just described, each 
daughter-nucleus, which now has the value of a germinal vesicle, 
first passes through the spirem stage of the metaphasis and 
then enters upon the stage of synapsis, namely, the nucleolus 
has a more or less central position, and all the chromatin of the 
nucleus becomes grouped immediately around it (Figs. 264-266, 
270, 271, 278), the peripheral part of the nucleus being trans- 
versed by only a few fine, unstaining strands of substance 
(linin ?). All intermediate grades between this and the preced¬ 
ing stage of the nucleus may be found. This is not an artificial 
appearance caused by the use of a particular preservative, since 
it is equally demonstrable on preparations fixed with aqueous or 
alcoholic corrosive sublimate, sublimate with acetic acid, Flem¬ 
ming’s fluid, and alcoholic solution of picric acid; only after 
the use of Perenyi’s fluid is this arrangement of the chromatin 
not found, but this fluid seems to be rather a poor one for most 
cytological study. It cannot be an artefact, since this appear¬ 
ance is found only in ova of a certain size but not in those 
which are larger ; thus it cannot be produced by the resistance 
offered by the cell membrane to the penetration of the fixa¬ 
tives, since this membrane is much thicker in the larger ova. 
This central arrangement of the chromatin then represents a 
definite stage of the germinal vesicle concomitant with the 
first appearance of yolk globules in the cytoplasm. 1 So at this 
point we may briefly describe the yolk development and then 
return to the changes of the nucleolus. 

The yolk first arises in the cell during the stage just 
described, that is, immediately after the conclusion of the spirem 
stage of the metaphasis. It appears in the form of small glob¬ 
ules ( Yk . 67 ., Figs. 262-264, 266, 270, 271), most of which are 
arranged close to the outer surface of the nuclear membrane, 
the first globules rarely arising at a distance from the nucleus. 
At this period they stain less deeply than later. The yolk 

1 This stage of synapsis (Moore) appears to be characteristic of the anaphase 
of the last spermatogonic and spermatocytic division in all the higher animals, 
and no doubt can any longer be expressed of its representing an artefact, 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


rapidly increases in amount, spreading from the region of the 
nucleus (which is central) to the cell periphery. In the largest 
ovarial ova the cytoplasm is densely filled with larger and 
smaller yolk globules; the larger ones appear homogeneous 
when stained with eosin (Fig. 269), but the Ehrlich-Biondi stain 
shows them to be composite masses of small globules. 

The nucleolus rapidly increases in size, at a somewhat greater 
proportionate rate than the nucleus itself. It is now large 
enough for its structure to be clearly made out : it consists of 
a homogeneous ground substance, which seems to stain more 
deeply with eosin as it grows larger ; a limiting membrane is 
clearly demonstrable in the largest nucleoli (Figs. 271-277, 
279-281) after staining by the Ehrlich-Biondi method or 
after fixation with Flemming’s fluid, though it does not differ 
chemically or in structure from the ground substance and is 
only a thin layer of the latter in which vacuoles never occur. 
At the close of the metaphasis of the mitosis small vacuoles 
make their first appearance in the ground substance of the 
nucleolus (Figs. 263 and 270). There are only a few of them at 
the start, but their number rapidly increases as the nucleolus 
grows larger, until there are large numbers of them in its center 
(Figs. 268 and 269). They are always more numerous at the 
center than at the periphery of the nucleolus, and usually first 
appear at the former point. On preparations stained with 
eosin the small vacuoles appear either as clear spaces or as 
black granules, according to the focusing of the microscope ; 
after the use of the Ehrlich-Biondi stain they become a light 
grayish color (note the contrast, — that in the eggs of Doto and 
Montagua the nucleoli appear as black granules only after 
treatment with the latter stain) ; after fixation in the fluid of 
Flemming the substance of these vacuoles is of a lighter color 
than the ground substance. This vacuolar substance is homo¬ 
geneous, and is probably of a thin, fluid nature. With the 
growth of the nucleolus the number of the vacuoles becomes 
very great, though their size does not seem to increase. In the 
nucleoli of the largest germinal vesicles examined the vacuoles 
no longer retain their original spherical form, but become mutu¬ 
ally confluent to some degree, not in such a manner as to pro- 


460 


MONTGOMERY. 


[Vol. XV. 


duce one or a few large vacuoles, but as to produce an irregular 
canicular network of vacuolar substance in the nucleolus (Figs. 
272-277, 279-281). This process often goes so far that in the 
largest nucleoli the deeply staining ground substance may 
appear in the form of a skein of threads, or merely of scattered 
granules surrounded by vacuolar substance. Especially on 
preparations stained by the Ehrlich-Biondi method is the skein¬ 
like arrangement of the ground substance well marked. I 
have no doubt that it was the observation of similar nucleoli in 
like stages which led Carnoy to the assumption of a “ nucleole- 
noyau,” that is, a nucleolus with a limiting membrane, and 
containing a wound thread of chromatin ; it is probable that 
Carnoy mistook the reticulum of the true ground substance of 
the nucleolus for chromatin, and considered what is really vacuo¬ 
lar substance to be the original ground substance; only 
studies on the genesis of a nucleolus can explain its various 
components. 

In the largest ova found in the body cavity the nucleolus 
reaches its maximum size (Figs. 279-281). It contains a greater 
amount of vacuolar than of ground substance, and instead of 
being regularly oval, as it was before, is often quite irregular 
in form, and very frequently apposed to the nuclear membrane 
(a position not noticed in any of the preceding stages). Whether 
this irregularity of form denotes the commencement of a degen¬ 
eration of the nucleolus I cannot say, since I had no prepara¬ 
tions of the stages of reduction. 

Two nucleoli were found in only two germinal vesicles (Figs. 
262 and 266), and in a spirem stage of an ovogonium three small 
nucleoli were present in one nucleus (Fig. 261). In the hun¬ 
dreds of other resting nuclei examined a single nucleolus was 
invariably present. These exceptional cases must, therefore, 
be considered abnormal, and not typical for certain stages of 
the nucleus. 

In the larger germinal vesicles there is a peculiar body in 
contact with the nucleolus, which remains to be described. 
This body (nx., Figs. 272, 274-277, 279, 281) is homogeneous, 
somewhat refractive, and lies either in close contact with the 
surface of the nucleolus, projecting beyond the periphery of the 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 461 


latter, or (and this is the rule for the largest, irregular nucleoli) 
it is imbedded in the peripheral portion of the nucleolus ; in 
the former position it is concavo-convex, in the latter, bicon¬ 
vex in outline, always being thickest in its median diameter. 
With the Ehrlich-Biondi staining method it almost invariably 
colors yellowish, and in only one or two cases did it stain 
somewhat similarly to the ground substance of the nucleolus ; 
after fixation in Flemming’s fluid, and staining with safranin, 
gentian violet, and orange G., it always appeared yellowish, 
while the ground substance remained wholly unstained. The 
largest nucleoli, i.e those of the largest germinal vesicles, 
have always at least one of these bodies in contact with their 
surface, but quite frequently two may be found on opposite 
sides of the nucleolus, and in one case I found three (Fig. 277). 
Those of different nucleoli vary slightly in their dimensions, 
but my observations give no clue as to their origin. All that 
can be said of their growth is that in the smaller nucleoli they 
lie upon the surface of the latter, while they are sunk into the 
peripheral portion of the larger nucleoli. It differs both chem¬ 
ically and structurally from the ground substance of the nucleo¬ 
lus, and from the vacuolar substance ; and it would seem to be 
derived from some part of the nucleus outside of the nucleolus, 
since it at first lies upon the surface of the nucleolus. This 
body may be comparable to the “ Nebennucleolus ” described 
by Flemming in the egg of Anodonta ; but I have found no 
structure in any of the other ova here examined which is 
identical with it. 

Yolk globules are assimilated by the nucleus from the cyto¬ 
plasm, though without the production of amoeboid processes. 
Such assimilated globules are usually of small size, but some¬ 
times large, compound ones are taken into the nucleus (Figs. 
267-269, 272, 274, 280); they occur most frequently singly or 
in small masses close to the inner surface of the nuclear mem¬ 
brane (Figs. 274 and 280) in almost all of the larger germinal 
vesicles, and in a few cases some globules may be found near 
the center of the nucleus. Careful observation shows that the 
yolk globules really occur within the nucleus, and are not arti¬ 
ficially removed there by the knife in sectioning. Usually these 


462 


MONTGOMERY. 


[Vol. XV. 


stain in the same manner as those contained in the cytoplasm. 
But occasionally from one to three of the larger globules (Fig. 
267) in the nucleus stain much more intensely than the others, 
though intermediate degrees of staining are to be found between 
these largest, most deeply colored ones and the smaller, less 
deeply stained ones ; so that there can be no doubt of the 
genetic relation of the two kinds. By staining with eosin 
these largest yolk globules in the nucleus stain almost or quite 
as deeply as the nucleolus itself, so that at first I mistook them 
for nucleoli ; but that they are chemically metamorphosed yolk 
globules and not nucleoli is shown, even leaving aside the fact 
that all intermediate forms may be found between them and 
the less deeply staining globules of the cytoplasm, by the fact 
that vacuoles are never found within them. By the Ehrlich- 
Biondi staining method no color differentiation was to be 
obtained for the larger and smaller yolk globules of the nucleus. 
But nevertheless I would think that these large yolk globules 
(or accumulations of such globules) which have been taken 
into the nucleus from the cytoplasm and there have undergone 
some degree of chemical change, possibly stand in genetic 
connection with that body which is apposed to the nucleolus 
in the larger germinal vesicles, and which has been described 
in the preceding paragraph. 

Chromatin. —We found the chromatin in the primitive peri¬ 
toneal cells and in the youngest ovogonia to be arranged in the 
form of granules (Figs. 250-254). In the following mitoses it 
is arranged in the form of a spirem, then of chromosomes, and 
again of a spirem (Figs. 255-261). Just after the conclusion 
of the spirem stage (of the metaphasis) it comes to lie in a 
more or less dense mass around the nucleolus, this mass 
appearing to be composed of a reticulum of short fibers (Figs. 
263-266, 270, 271, 278). In all these stages the chromatin is 
marked by its deep blue staining with haematoxylin. After 
the last stage described it gradually departs from the close 
vicinity of the nucleolus and becomes evenly distributed 
throughout the nucleus. But when it has thus become diffused 
it does not stain with haematoxylin as before, but appears in 
the form of a very large number of minute microsomes, which 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 463 

appear not to stain at all, and of a few delicate fibers, which 
stain a lilac color (Figs. 267-269). As the germinal vesicle 
increases in size these chromatin fibers gradually become 
thicker and more numerous, commence to stain more deeply 
with haematoxylin, and gradually connect together to build 
a chromatin reticulum ; the minute, unstained microsomes 
still occur between these fibers. Finally, in the largest nuclei 
at my command, and ones which had been fixed with the fluid of 
Flemming and stained by the triple stain of this cytologist, we 
find, in addition to the abundant unstained microsomes, short, 
rod-like masses of chromatin, which stain deeply with gentian 
violet, and each appears to be formed of a row of granules or 
thickened discs (Fig. 280). Whether the minute microsomes 
are true chromatin or are lanthanin (oedematin) granules is 
open to question ; the latter assumption might be the correct 
one. We notice two remarkable phenomena in the chromatin 
changes just depicted : (1) the grouping of the chromatin in 
the center of the nucleus, around the nucleolus, at the comple¬ 
tion of the mitotic stages ; and (2) immediately subsequent to 
the preceding, the lilac stain of the chromatin after haematoxy¬ 
lin. Now, concomitant with the former of these two phenomena, 
the yolk makes its first appearance in the cytoplasm, and as we 
have shown above, usually in the close vicinity of the nucleus. 
It would be quite erroneous to conclude that the yolk globules 
are in any way produced by the chromatin, as e.g. y by a migra¬ 
tion of chromatin particles out of the nucleus ; for in this stage 
all the chromatin is removed from the periphery of the nucleus. 
On the other hand, however, I would suggest the hypothesis 
that the reason for the chromatin being removed from the 
periphery of the nucleus is because at this period the peripheral 
portion of the latter is chiefly concerned in the assimilation of 
yolk substance from the cytoplasm. In support of this assump¬ 
tion the fact may be recalled that in the following stage the 
chromatin fibers are stained a lilac color, as if they were stained 
with eosin, as well as haematoxylin, and not as before, simply 
with the former stain ; this would show that during this period 
there is an addition of a cytoplasmic substance to the chromatin 
fibers, perhaps allied to the substance of the yolk globules, and 


464 


MONTGOMERY. 


[Vol. XV. 


this substance would superinduce the lilac staining of the 
chromatin threads. This addition of a probably nutritive 
substance would seem necessary, in order that the amount of 
the chromatin continue to increase as the nucleus itself grows 
larger. Subsequently all that nutritive substance attached to 
the chromatin threads would seem to become metamorphosed 
into chromatin, since in the largest germinal vesicles these 
threads again stain a deep blue. And as a matter of fact, the 
quantity of the chromatin must increase with the growth of the 
ovum, since it can easily be demonstrated that in the larger 
nuclei there is an absolutely greater amount of this chromatin 
present than in the nuclei of the primitive peritoneal cells. 1 

ii. Piscicola rapax (Verr.) (= Pontobdella rapax Verr., which 
Dr. Percy J. Moore assures me is a true Piscicola). 

(Plate 29 , Figs. 300 - 316 .) 

(The ovary is a tubular, contorted sack; from its inner sur¬ 
face numerous smaller, likewise tubular (round on cross-section), 
acini project into its cavity, each acinus containing numerous 
ovogenetic stages, the least mature of which lie at its proximal 
end, the most mature at its distal. These several acini are 
not continued as far as the external opening of the ovary, but 
their distal ends apparently open into a large ovarial cavity, 
and the ova drop into this cavity before they can arrive at the 
external genital opening. Each single acinus of this leech may 
be compared to either of the two whole ovaries of Ascaris.) 

The youngest ovarial stages are small ovogonia in stages of 
mitotic division (Fig. 300). In them no nucleoli were to be 
seen; a minute nucleolus might be present in each of these 
nuclei and be obscured by the dense mass of chromatin. In 
all stages subsequent to these a single nucleolus is present in 
the nucleus (now a germinal vesicle) until the pole spindle is 
formed; in the smaller nuclei the nucleolus is usually oval, in 
the larger ones spherical. The growth of the nucleolus keeps 

1 For the researches of other authors on germinal spots of polychaetous anne¬ 
lids, cf. the reviews of the papers of Korschelt (’89, ’95), Graff (’88), Giard (’81), 
Vejdovsky (’82), Eisig (’87), Fraipont (’87), Mead (’95), Fauvel (’97), Michel 
(’96), and Carnoy (’84). 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES . 465 

pace proportionally to that of the nucleus (Figs. 301-304). 
Then vacuoles arise in the nucleolus, these being neither very 
numerous nor very minute (Figs. 304-310, 312-316). The time 
when these vacuoles first arise is very variable, though in the 
majority of cases they do not appear before the nuclear sap 
begins to stain red. The size of the nucleolus does not 
always stand in the same proportion to that of the nucleus. 
Its ground substance is dense, stains deeply with eosin, and 
no limiting membrane is present; but by the use of the double 
stain Lyons blue and acid carmine, whereby the nucleolus 
stains blue and the chromatin red, a deep red line appears to 
surround the nucleolus: I cannot determine whether this 
line is a nucleolar membrane or a layer of chromatin, or 
whether it is merely an appearance caused by the refraction 
of the nucleolus. 

When the nucleolus first appears it is usually situated at 
that pole of the nucleus opposite the chromatin mass and is 
not in contact with the nuclear membrane (Fig. 301). In 
nuclei of intermediate size, before the nuclear sap commences 
to stain with eosin, it is most frequently in contact with the 
nuclear membrane (Figs. 302-304); but in the largest ger¬ 
minal vesicles it is never in contact with this membrane, 
though often lying excentrically in the nucleus. 

As soon as the germinal vesicle has nearly, or quite, attained 
its maximum dimensions (quite frequently, however, in those of 
still smaller size) two very noticeable changes take place in it: 

(1) the chromatin assumes a different form and stains differ¬ 
ently (these chromatin changes shall be delineated later); and 

(2) the nuclear sap, which had heretofore remained colorless or 
was merely of a light lilac shade (by the double stain haema- 
toxylin and eosin), now becomes a yellowish-red color, so that 
the nuclei in this stage may be easily distinguished from those 
of preceding ones (Figs. 304, 305, 307-310, 316). Simul¬ 
taneously two changes occur in the nucleolus: (1) it stains no 
longer a deep red with eosin, as before, but a yellowish red, 
and appears more refractive; and (2) the several vacuoles 
within it gradually fuse together and so produce a larger one, 
which has usually a central position. The fluid, structureless 


466 


MONTGOMER V. 


[Vol. XV. 


substance of the vacuole stains more faintly than the ground 
substance of the nucleolus, and has much the same color shade 
as the nuclear sap. In certain germinal vesicles, which appear 
to be of a somewhat later stage of development, numerous 
small globules (j n.D ., Figs. 306 and 310) are scattered through 
the nuclear sap; they stain with eosin a little more deeply than 
the last-named nuclear portion, vary in number and size, and 
have no regular distribution. In one case (Fig. 316), which 
stood in a stage immediately antecedent to the pole spindle 
formation, where there was a centrosome at either end of the 
nucleus in the cytoplasm (the nuclear membrane had not yet 
disappeared), such globules were present in the nucleus; so 
that we may infer that these globules are one of the latest 
formations in the germinal vesicle before the pole spindle is 
formed. I have not found any stages between the stage just 
described and the perfectly formed spindle (Fig. 311). About 
fifty or sixty ova were examined in the stage of the first pole 
spindle, and in all of them the nucleolus had completely 
disappeared, and no trace of it could be found either in the 
nucleus or in the cytoplasm. 

What has been the manner of this disappearance of the 
nucleolus ? Its total disappearance must occur within a rela¬ 
tively short time, since otherwise one would expect to find 
stages showing this process. The observations which I was 
able to make would demonstrate at least the mode of the com¬ 
mencement of the vanishing of the nucleolus. We have seen 
that when the germinal vesicle has attained its greatest size 
or, in some cases, a little before its maximum size is reached, 
its nuclear sap stains red; therefore some substance must be 
suspended in the caryolymph at this period which was not con¬ 
tained in it before. Now such a substance must have been 
derived either from other elements of the nucleus or from 
the cytoplasm. It has probably not been derived from the 
cytoplasm, since the nuclear membrane at this stage has its 
maximum thickness and hence could not be easily penetrable; 
and also there is no appearance of any similar substance in the 
cytoplasm, since no yolk globules or other nutritive elements 
seem to be present, but the whole cytoplasm (at least the 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 467 


nodules of its meshes) stains a lilac-blue color. And since 
it is wholly improbable that it should be derived from the 
chromatin we must conclude that it takes its origin from 
the nucleolus. In other words, a substance emanates from the> 
nucleolus and dissolves in the nuclear sap, and this process 
must be regarded as the commencement of the dissolution of 
the nucleolus. In support of this conclusion is the fact that 
in many germinal vesicles the nuclear sap stains most intensely 
in the neighborhood of the nucleolus (Fig. 309). Further, the 
minute red-staining globules which later occur in the nuclear 
sap must also be nucleolar in point of formation, i.e., be either 
a substance given off in globular form from the nucleolus, or 
be accumulations (perhaps chemically changed by the action of 
the nuclear sap) of that nucleolar substance which has already 
diffused through the nucleus. Of importance in this connec¬ 
tion is the fact that these globules are often found in contact 
with the nucleolus (Figs. 306 and 316). In all preceding stages 
the nucleolus is regularly oval or spherical in outline, but in 
the largest germinal vesicles not only may the size of its con¬ 
tained vacuole be increased to such an extent that the original 
ground substance forms only a thin shell around it (Figs. 308, 
312, 314), but also its outline becomes frequently irregular 
(Fig. 313); and in one case I found it broken at one pole, so 
that its large vacuole communicated with the cavity of the 
nucleus (Fig. 315). A morphological change in the shape of 
the nucleolus which seems to take place with great regularity 
consists in the indentation of the nucleolar wall at that point 
where it is thinnest (Figs. 308, 314, 316). It would seem that 
the pressure from without, i.e., the pressure of the nuclear sap, 
being greater than the pressure of the fluid within the vacuole, 
would cause the nucleolar wall to be pressed in at that point 
where it is thinnest. The fact remains that the nucleolus 
persists in the nucleus until a very short time before the pro¬ 
duction of the pole spindle, and when the latter is formed no 
trace of it can longer be found in any part of the nucleus or 
cell. And since there is no reason for supposing that it is 
extruded from the cell we must assume that it dissolves 
within it. The red-stained substance and small globules 


468 


MONTGOMERY. 


[Vol. XV. 


within the nucleus would show that dissolution commences 
in the nucleus; and we must assume that when the nuclear 
membrane has disappeared the cytoplasmic substances which 
then come into contact with the nucleolus would cause its 
rapid and total dissolution. It may be remarked that in the 
region of the fully formed spindle (Fig. 311) no trace of the 
red-stained nuclear sap is longer to be seen; accordingly this 
sap with its contained nucleolar substance must either have 
been distributed through the cytoplasm or have been chem¬ 
ically changed by that portion of the latter which immediately 
surrounds the spindle. 

In the ovary no ova are to be found which have advanced 
beyond the production of the first pole body, so that the forma¬ 
tion of the second pole body must occur after the egg has been 
discharged from the ovary; I had no material at hand to enable 
me to determine the relation of the nucleolar substance in the 
female pronucleus. 

Of considerable morphological interest are the metamor¬ 
phoses of the chromatin in the various ovarial stages. In those 
small ovogonic mitoses (Fig. 300) from which the true egg cells 
(first ovocytes) are derived aster and dyaster stages are to be 
found ; with the lens used for this study (the homogeneous 
immersion ^ of Zeiss) I could not determine the form of the 
chromosomes. As the ovum increases in size the dense chro¬ 
matin mass of the aster gradually loosens, until up to the time 
when the nuclear sap commences to stain red (Figs. 301-304) 
the chromatin is arranged in the form of rather numerous 
granules, which are situated mostly close to the nuclear mem¬ 
brane. Thus far the chromatin has stained intensely blue, with 
the double stain haematoxylin and eosin ; but when the nuclear 
sap begins to stain with eosin a marked change takes place in 
the character and arrangement of the chromatin; it now stains 
a lilac color, often more reddish than bluish, and has no longer 
a peripheral position, but becomes arranged in the form of 
threads, sometimes in the form of a small number of loops, the 
two ends of each loop being joined together (Figs. 304, 305, 
307) 3°9)- In some of the larger germinal vesicles absolutely 
no trace of chromatin can be found (Fig. 316). In the equator 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES . 469 

of the first pole spindle (Fig. 311) lie twelve small chromosomes, 
which stain an intense blue black with haematoxylin and have 
an oval or slightly elongate form. It remains for investigators 
working with more abundant material and with stronger micro¬ 
scopical lenses, to penetrate more deeply into these phenomena 
of the chromatin changes, but it would seem that the chromo¬ 
somes of the first pole spindle have the value of either tetrads 
or dyads. The lilac or even reddish stain of the chromatin at 
a particular period would seem at first sight to be due to the 
assimilation by the chromatin of that nucleolar substance dif¬ 
fused in the nuclear sap ; but even as probably it might be due 
to the mere penetration of this substance between the individ¬ 
ual microsomes of each chromatin thread, without any chemical 
change of the chromatin substance (Fig. 309). The red- 
staining globules in the nuclear sap, which I have assumed 
to be of nucleolar derivation, cannot be considered as meta¬ 
morphosed portions of chromatin substance, since they vary 
so considerably in size and number ; this point needs to be 
emphasized, since in some of the larger germinal vesicles no 
trace of chromatin is to be seen, and it might be thought by 
some one that these globules, which occur in such nuclei, repre¬ 
sented the supposedly absent chromatin. (Platner, ’ 89 c, had, 
in Aulastommn seen only nucleolar fragments and overlooked 
the true chromosomes.) Where in the largest germinal vesi¬ 
cles, before the formation of the pole spindle, the chromatin 
appears to be absent in the nucleus, we must assume that it 
is merely obscured by the large amount of diffused nucleolar 
substance. 

In the first pole spindle (Fig. 311), after treatment with 
Flemming’s fluid or with corrosive sublimate, the mantle fibers 
have a remarkable thickness and appear even thicker than in 
Fig. 311 ; they stain a reddish-lilac color with the haematoxylin 
and eosin stain, not a lilac blue, as do the rays of the asters 
and the cytoplasm; I could not determine whether they extend 
quite to the centrosomes. I am also unable to decide whether 
each chromosome lies upon a single spindle fiber which extends 
from centrosome to centrosome, or whether its ends are con¬ 
nected with separate fibers. The centrosomes are rather large, 


470 


MONTGOMERY. 


[Vol. XV. 


refractive granules, and stain with eosin ; they were present in 
one egg, close to, and opposite, the two poles of the nucleus, 
before the nuclear membrane had disappeared (Fig. 316), so that 
they may be extranuclear in origin. The radiations of the asters 
are very clear, especially after fixation in Flemming’s fluid, and 
may be traced nearly to the cell membrane. Immediately 
around each centrosome a central portion of the aster is dif¬ 
ferentiated, namely, an attraction sphere (in the terminology of 
van Beneden), and this differs from the remaining portion in 
staining less intensely, and appears to be quite sharply bounded 
from it. In this attraction-sphere the cytoplasmic granules are 
smaller and more densely grouped, so that at first sight it 
might appear to consist of a homogeneous “ archoplasm,” but 
careful study shows that in it the cytoplasmic microsomes are 
arranged in radial rows around the centrosome, and each of 
these rows appears to be continuous with a ray of the outer 
aster. Or, to express it differently, the microsomic rays of the 
sphere extend to the centrosome, but this terminal part of each 
ray differs from the remaining distal portion in that its micro¬ 
somes are smaller and closer together. Thus in Piscicola the 
finer structure of the attraction-sphere seems to have much 
resemblance to that of Ascaris, as described by Kostanecki and 
Siedlecki {Arch. mikr. Anat ., 48, 1896). 

It remains to describe the mode of arrangement of the ova 
within each ovarial acinus. The proximal, small end of the latter 
is filled with small ovogonia (the youngest stages), and from 
mutual contact these are polygonal in form (Fig. 300). As we 
proceed towards the distal end of the acinus (Fig. 301) the ova 
not only become gradually larger, but have a different arrange¬ 
ment, in such a manner that they become epithelially grouped 
along the wall of the acinus, each cell having a pyramidal 
shape, with its apical end directed towards the central cavity 
of the acinus. A little more distally in the acinus (Figs. 302 
and 303), the ova become not only larger, but fewer of them 
are to be found on a given transverse section of the acinus ; the 
individual ova have more of an oval shape and become sepa¬ 
rated from one another. Now when we proceed still further 
towards the distal end of the acinus (Fig. 304) we find a single 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 471 


ovum commencing to outstrip the others in point of size, i.e. t 
in rapidity of growth, until we reach a point where this fortu¬ 
nate cell nearly fills the whole cavity of the acinus, driving the 
neighboring ova aside. Those cells which come into contact 
with such a rapidly growing ovum, as well as those in more 
proximal portions of the acinus which did not chance to lie 
close to the wall of the acinus, do not develop further, but 
disintegrate, and various stages of such disintegration may be 
seen in the cavity of the acinus, such as irregular cells with a 
nucleus, those which have lost their nuclei, and finally refrac¬ 
tive cytoplasmic masses which stain deeply with eosin (the 
cytoplasm of the developing ova stains with haematoxylin). 
Perhaps such degenerated masses of cellular substance are des¬ 
tined to be assimilated by their more fortunate brethren. Often 
a number of such degenerating ova are to be seen grouped at 
one pole of a large ovum, and these cases present a certain 
similarity to cleavage stages, the large ovum resembling a 
macromere, the others micromeres. It is not difficult to find 
an explanation for the disintegration of certain of the ova, for 
only those close to the wall of the acinus can procure nourish¬ 
ment in amount sufficient for their growth, since this nour¬ 
ishment must be derived through the wall of the acinus from 
the body cavity (there being no yolk in the ova); and the 
peripherally situated ova must obtain all the nourishment thus 
furnished, so that those in the center of the acinus simply die 
for want of food. Further, a particular ovum of those placed 
peripherally, if it procures a greater amount of nourishment 
than its neighbors do, because, e.g v of being in contact with a 
greater surface of the wall of the acinus, grows faster than the 
others and, pushing them aside, eventually gets full control of 
the whole amount of nourishment, so that a slight advantage at 
the start would increase in value in a geometrical ratio. Here, 
accordingly, we have a beautiful example of that process termed 
by Roux “der Kampf der Theile urns Dasein,” that cell becoming 
a mature ovum which has succeeded in obtaining the greatest 
amount of nourishment. It is also interesting to note the 
position of the nucleus within the growing ovum ; in all the 
younger stages of the egg it is placed in that part of the cell 


47 2 


MONTGOMERY. 


[Vol. XV. 


which is nearest to the wall of the acinus, i.e ., nearest to the 
source of the food supply ; only then does it come to occupy a 
central position within the cell, when the latter has attained its 
maximum size and the thickness of the cell membrane shows 
that the cell is assimilating little or no nourishment from 
without. 1 

b. Somatic Cells. 

12 . Ganglion Cells of Do to. 

(Plate 21 , Figs. 36 - 49 .) 

(I have studied those nerve cells which occur in the cerebral, 
pleural, and pedal ganglia. Three kinds of these cells may be 
readily distinguished and described in succession : (1) colossal 
cells, which are found only in the cerebral ganglion; (2) cells of 
medium size; and (3) small cells.) 

Colossal ganglion cells ( Figs. 43-49).—The number of the nu¬ 
cleoli in the nuclei of these cells varies from about six to thirteen; 
they are also variable in regard to the position which they 
occupy in the nucleus, and though always excentrically placed 
they never lie in contact with the nuclear membrane. Some¬ 
times all the nucleoli in a given nucleus are of approximately 
equal size, but as frequently one or two are several times larger 
than any of the others. Where such larger nucleoli occur along 
with a number of smaller ones, the former are usually vacuolar 
in structure ; sometimes nearly all the nucleoli contain vacuoles, 
in other cases none of them are vacuolar. Quite often the 
nucleoli in a nucleus show slight differences in their staining 
intensity, and one of them may stain quite differently from the 
rest (Figs. 44 and 46). None of the nucleoli have limiting 
membranes. No cases of nucleolar division were found, unless 
those cases where two nucleoli lie near to one another may 
represent the completion of such a division. 

Ganglion cells of medium size (Figs. 37-42). —In these the nu¬ 
cleoli vary in number from one to four, two or three being the rule. 
Those of the same nucleus frequently show differences in size 
and form, as well as slight staining differences. In only one 

1 For the observations of other writers on germinal spots in Hirudinea , cf. 
O. Hertwig (’76), Leydig (’49), Whitman (’78), and Platner (’89c). 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


case (Fig. 41) I found three nucleoli of approximately equal 
dimensions and homogeneous ; usually they vary somewhat in 
size and contain vacuoles. The shape of the nucleoli is either 
spherical or oval, or it may be irregular; certain ones stain 
scarcely at all, and appear granular : these might represent 
cases of degeneration. 

Smallest ganglion cells (Fig. 36). —Here a single nucleolus 
is the rule, though two may occasionally be found. They are 
spherical or oval, and vary considerably in size. Vacuoles do 
not seem to occur in them, though they might well escape 
observation from the small dimensions of the nucleoli, which 
often renders it difficult to distinguish the nucleoli from the 
larger chromatin granules. 

In all these ganglion cells the chromatin appears in the form 
of small granules, but on a preparation fixed with Hermann’s 
fluid and stained with Lyons blue (Fig. 45) it appeared as a 
network ; in this preparation the granules seemed to be united 
by fine fibers, which stained less intensely than the granules. 
But even here the connecting threads might consist rather 
of linin than of chromatin, since the solution of Lyons blue 
employed by me stained all the nuclear substances except the 
nuclear sap (paralinin). Such fibers often appear to radiate 
outwards from the surface of the nucleoli, as if the latter were 
suspended by them. The nucleoli always stain differently from 
the chromatin. 

There is, as a rule, a relatively small amount of nucleolar sub¬ 
stance in the cells of the second and third types in comparison 
with most of the other nuclei which I have examined ; but the 
nuclei of those of the first type, on the contrary, usually contain 
a relatively large amount of this substance, for not only may 
one or two of the nucleoli in a nucleus be quite large, but also 
a considerable number of nucleoli are frequently present. 


13. Ganglion Cells of Montagua fiilata (Verr.). 

(Plate 22 , Figs. 90 - 97 .) 

(The same types of cells may be roughly distinguished as in 
Do to.) 


474 


MONTGOMERY. 


[Vol. XV. 


Colossal ganglion cells (Figs. 90-92, 94-97). —In the nuclei of 
these there are never more than from one to three nucleoli, 
which neither contain vacuoles nor become noticeably irregular 
in size, as is the case in Doto. Most frequently only a single 
nucleolus is present. It is the rule that they are oval and not 
spherical, though in some cases they may appear perfectly 
spherical ; perhaps the great majority of them are oval and 
seem to be spherical only when they do not chance to be longi¬ 
tudinally sectioned. Their substance is perfectly homogeneous, 
without a limiting membrane. When two or three occur in the 
same nucleus they are usually of approximately equal dimen¬ 
sions (Figs. 94 and 95). Further, it would seem to be the rule 
that when one nucleolus is present in a nucleus it is larger than 
any one of the two or three which may be found in other 
nuclei ; but, nevertheless, the relative amount of nucleolar 
substance seems to vary in different nuclei. 

Ganglion cells of medium size (Fig. 93). — Here one or two 
nucleoli are present in each nucleus, and these are of homo¬ 
geneous appearance. 

Smallest ganglion cells. — The nucleoli are similar to those 
of the corresponding cells of Doto. 

On a preparation preserved in Flemming’s fluid I find many 
of the nucleoli present a different structure from those fixed 
with corrosive sublimate or Kleinenberg’s fluid. Thus many 
of them do not appear homogeneous, but finely granular and 
refractive (Figs. 96 and 97). On the surface of such nucleoli 
occur small, refractive, yellowish globules, which appear black or 
yellow, according to the focus of the microscope; some of them 
are very small. These never occur within the nucleolus, but 
only on its periphery. They may easily be distinguished from 
the chromatin granules by their rounded form and high degree 
of refrangibility, as well as by their deeper yellow color (this 
preparation had been stained with haematoxylin and eosin, but 
the nuclei had not become stained, probably owing to too long 
a fixation in the fluid of Flemming). Numerous other nuclei 
on the same sections showed none of these globules, and none 
were to be seen on preparations which had been differently 
preserved. Accordingly, I consider them to be artefacts. 


Nro. 2.] COMPARATIVE CVTOLOGICAL STUDIES . 


caused by (i) the direct action of the fluid of Flemming, or 
more probably (2) they might be post-mortem exudations of the 
nucleoli, which might well be produced before the slowly pene¬ 
trating fixative had reached to the cells in question. At any 
rate, they cannot be regarded as normal structures. Do they 
represent the “ Kernkorperchenkreis ” of Eimer ? 

The chromatin, as in Doto , occurs in the form of granules, 
which are connected by fine fibers. After fixation with Klein- 
enberg’s fluid a clear space encloses each nucleolus (Figs. 93 
and 94); but this space is not to be found after fixation in 
other fluids. 

As in Doto , the nuclei of the colossal ganglion cells contain 
a relatively greater amount of nucleolar substance than do those 
of the second and third types. But in the former genus there are 
in the colossal cells from about six to thirteen nucleoli, and these 
vary noticeably in size and structure, while in Montagna there 
are only from one to three, which are always homogeneous and 
usually quite equal in dimensions. Why should there be this 
marked difference in the form and number of the nucleoli ? 1 

14. Ganglion Cells of Piscico la rap ax (Verr.). 

(Plate 23, Figs. 134-136 ) 

In the ganglia of the brain occur cells of different dimen¬ 
sions. Each nucleus contains most usually a single small 
spherical nucleolus; seldom are there two present, and in these 
cases they are unequal in size. None of the nucleoli contain 
vacuoles. They are excentric in position, but are never in 
contact with the nuclear membrane. These nucleoli are small 
in proportion to the size of the nucleus. 

15. Muscle Cells of Linens gesserensis (O. F. M.). 

(Plate 21 , Figs. 51 - 56 .) 

(The nuclei of the circular muscular layer of the body wall 
were studied. Those of Cerebratulus lacteus Verr. are essen- 

1 For other observations on nucleoli in ganglion cells of molluscs, cf the 
reviews of the papers of Pfliicke (’95), Leydig (’83), and Rohde (96). 


476 


MONTGOMERY. 


[Vol. XV. 


tially similar to those of Linens ; in the metanemerteans they 
are too small for satisfactory study.) 

These nuclei are very variable in shape, all extremes being 
found between ovoid or oval and elongate rod-like forms. But 
they are rarely angular. I have remarked in a previous con¬ 
tribution that the nuclei of the muscle cells are more variable 
in form than those of the cells of any other tissue in the 
nemerteans, and now I would offer the following explanation 
for this variability : when the muscle fiber (a single, smooth 
fiber with attached nucleus constitutes a muscle cell) contracts, 
this contraction must produce likewise a contraction (shorten¬ 
ing) of the nucleus ; but when the fiber expands the form 
of the nucleus must become more elongate, corresponding to 
the elastic extension of the fiber, for the fiber cannot contract 
without causing a shortening of its nucleus, since the latter is 
closely adherent to it. 

One very small nucleolus is usually to be seen in each 
nucleus (Figs. 51-54, 56); sometimes it does not appear to be 
present (Fig. 55), but whether in these cases it is absent or only 
escapes observation by reason of its minute size, it is difficult 
to decide; in the greater number of nuclei it may be seen by 
careful focussing of the microscope. It most usually lies very 
close to the center of the mass of nucleoplasm, so that if the 
nucleus be larger at one pole than at the other it is situated in 
the larger end, while in elongate nuclei, of nearly equal diam¬ 
eter throughout, it usually lies at an early equal distance from 
both ends of the nucleus. The nucleolus may be said, as a 
general rule, to occupy the center of the nuclear substance, 
and is not often markedly excentric; in none of the other cells 
examined in the course of these investigations did the nucleoli 
show a similar tendency to occupy the center of the nucleus. 
The nucleolus always stains differently from the chromatin. 

The relative amount of chromatin varies in different nuclei. 
It is always found, after the action of various fixatives, to occur 
in the form of small granules, which are connected by delicate 
irregular fibers, which stain exactly as the granules do. The 
nuclear sap stains faintly with haematoxylin (this has not 
been shown in the figures). The nucleolus is either in contact 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


with chromatin granules or with fibers of chromatin, which 
pass between it and the nuclear membrane; there is never a 
clear space around the nucleolus, but it seems to be held in 
position by the chromatin. 


16. Muscle Cells of Piscicola rapax (Verr.). 

(Plate 29, Figs. 325-337.) 

(The nuclei of the longitudinal muscle layer of the body wall 
were studied. For the examination of the different stages of 
these nuclei worms of different sizes must be studied; I exam¬ 
ined the nuclei of leeches of about 6 mm. in length, where the 
cells and their nuclei are smallest, as well as of larger and fully 
mature individuals, where these cells and their nuclei attain 
their maximum dimensions.) 

In the smallest nuclei (Fig. 325) a single nucleolus is inva¬ 
riably present and lies centrally ; it is of medium size, more or 
less oval in outline, and contains a varying number of small 
vacuoles. In larger nuclei it becomes larger and more elongate 
in form, lying in the longitudinal axis of the nucleus (Figs. 328 
and 331) ; at the end of this stage its greatest dimensions are 
reached. Next commences a process of fragmentation of this 
original nucleolus into a number of smaller nucleoli, which are 
of different sizes. There appears to be little uniformity in the 
mode of this nucleolar division (Figs. 327, 329, 332, 333): the 
nucleolus may become dumbbell shaped and then divide into 
two larger pieces; or when much elongated it usually breaks 
simultaneously into a number of consecutive portions ; or buds 
of nucleolar substance may be divided off from its surface. This 
segmentation is not strictly dependent upon the size of the 
nucleus, nor upon the size or form of the nucleolus. The frag¬ 
mentation continues, the larger daughter-nucleoli also dividing, 
until in the largest nuclei (those of the mature worm) as many 
as twelve small nucleoli may be present, which are irregularly 
distributed through the nucleus (Figs. 335 - 337 )- In a11 these 
stages at least some of the nucleoli contain vacuoles, though 
they have not been reproduced in all the corresponding figures. 

All the nucleoli of the largest nuclei are thus produced by a 


478 


MONTGOMERY. 


[Vol. XV. 


series of divisions from the single original one. This division 
usually commences, then, when the form of the nucleus changes 
from the original oval to a more elongate shape. It seems 
probable that this elongation of the nucleus may directly cause 
the division of the nucleolus, since the long axis of the latter 
coincides with that of the nucleus; and were the nucleolus in 
any way fixed in position in the nucleus, the nuclear elongation 
would draw out the nucleolus and cause it to break into frag¬ 
ments. But the division of the daughter-nucleoli does not 
always take place in the direction of the long axis of the 
nucleus, so that "some other factor might be at work to produce 
this division. 

The chromatin is arranged in the form of a reticulation (Fig. 
326). The nuclei of the younger cells are usually regular in 
outline, but those of the larger ones become very irregular; 
this irregularity of the contours of the nuclei is more marked 
by fixation with corrosive sublimate than with Flemming’s fluid, 
so that it might be regarded as an artefact caused, eg ., by the 
obstacle offered to the rapid penetration of the preserving 
fluid by the dense outer (fibrillar) layer of the cytoplasm in 
the largest muscle cells. 

17. Blood Corpuscles of Doto. 

(Plate 22, Figs. 98-101 ; Plate 23, Fig. 102.) 

(These cells are usually to be found abundantly in the 
cavity of the cirratida and of the sheaths of the tentacles, 
though their number varies greatly in different cirratida. 
They lie in the meshes of the loose network of mesenchym 
cells, either singly or grouped together into bundles. I have 
been unable to find them in other parts of the body. These 
cells appear to be free mesenchym cells, with perhaps the 
function of blood corpuscles.) 

There is always a single large nucleolus, which is usually 
very large in proportion to the size of the nucleus. It varies 
in form from a perfect sphere to an elongate oval. The nucleo¬ 
lar substance is usually homogeneous, but in some cases it is 
granular (Figs. 99-102) and then it stains faintly as if it were 


No. 2.] COMPARATIVE CVTOLOGICAL STUDIES. 


undergoing a degeneration. Quite frequently a small spherical 
granule lies in the center of the nucleolus and this always 
stains more intensely than the surrounding substance (Figs. 
99-102). In only about half a dozen cases, out of hundreds of 
cells examined, did I find attached to the surface of the nucleo¬ 
lus one or two much smaller bodies, which also stained less 
intensely (Figs. 100 and 101). Can it be that in certain cases 
the nucleolus becomes differentiated into a “ Hauptnucleolus ” 
and a “ Nebennucleolus,” in which case these small bodies 
would represent the “ Nebennucleoli ” ? In certain of the cir- 
ratida of a young individual the nucleoli of the greater num¬ 
ber of nuclei were situated at that pole of the nucleus directed 
towards the median axis of the cirratidum, i.e. f those in the 
nuclei on the right side of the cirratidum were in the left-hand 
poles of the nuclei, and those in the nuclei of the left side of 
the cirratidum (as seen on sections) were placed at the right- 
hand poles of the nuclei. I did not observe this regular posi¬ 
tion of the nucleoli in the cirratida of the other individuals 
sectioned and hence would conclude that it was not a normal 
phenomenon, but an osmotic consequent of the fixing reagent. 1 
The size of the nucleolus preserves approximately the same 
ratio to that of the nucleus. 

The nucleus is either spherical or oval in outline. The 
apparent arrangement of the chromatin varies according to the 
fixative employed. After picro-nitro-osmic acid (Fig. 102) it 
appears granular; after Hermann’s fluid (Figs. 99-101), in the 
form of delicate fibers % which radiate from the nucleolus to the 
nuclear membrane; after alcoholic solution of corrosive subli¬ 
mate (Fig. 98) we find a few fiber's radiating from the surface 
of the nucleolus, but the greater amount of the chromatin 
appears in the form of granular masses, which lie mainly near 

1 In a previous paper (’ 96 ) I figured for the nuclei of those mesenchym cells 
which surround the distal end of the ventral nerve cords of Cerebratulus lacteus, 
the nuclei with their chromatic masses pressed against that side of the nuclear 
membrane which was situated nearest to the central point of the section. At 
that time, I regarded this excentric position of the chromatin as a normal but 
peculiar phenomenon; but now, on comparison with the cells of Doto, I am 
convinced that it is an artefact produced by the osmotic action of the fixing 
reagent. 


480 


MONTGOMERY. 


[Vol. XV. 


the periphery of the cell, so that the nucleolus is surrounded 
by a clear space. These nuclei thus offer a suggestive object 
lesson, to teach how careful one must be in the determination 
of the form of delicate cellular structures by the study of 
preserved material. 

Cells which are isolated have a spherical form ; those grouped 
together are polygonal, owing to their mutual pressure (Figs. 
99 and 101). A cell membrane is present. The cytoplasm is 
for the most part finely granular; portions of it, however, are 
always more dense and stain more deeply than the former 
portion; there are great individual differences in different cells 
(Figs. 100 and 102). Often the cytoplasm is more or less vacuo¬ 
lar or a clear space may partially surround the nucleus and 
a similar space be present between the cytoplasm and the cell 
membrane, this space being transversed by a few radiating 
fibers. Such spaces are best shown after the action of the 
fluid of Hermann; they are seldom to be seen after fixation in 
picro-nitro-osmic acid ; but whether a coarse alveolar layer of 
cytoplasm at the periphery of the cell be normal or be an arte¬ 
fact, there are certainly marked differences in the structure of 
the cytoplasm in neighboring cells, and these differences might 
be regarded as the morphological changes corresponding to 
functional phases in the cells. Cases of degenerating cells are 
numerous, and may be recognized by their faint staining 
properties and by their granular appearance. 

18. Giant Cells of Dcto. 

(Plate 30.) 

(These enormous cells, which are the largest cells in the 
body, not excepting the ova, lie at the anterior part of the 
body just behind the head region and are closely apposed to 
the folds of the nidamental gland. They do not produce a 
closed mantle on the outer surface of this gland, but either are 
isolated or occur in small groups of from two to four cells 
each. In each individual their number appears to be about 
thirty or forty. These cells do not seem to have any open 
communication with the neighboring tissues, and I cannot 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 481 


conclude from their structure what their function is; per¬ 
haps they have a function similar to that of lymph glands. 
Such cells are absent in Montagna.) 

The form of these is a more or less polyhedral one, caused 
by the pressure of the surrounding organs (Fig. 339). The 
nucleus is relatively and absolutely very large and is very vari¬ 
able in form, sometimes irregularly oval, sometimes with obtuse 
or pointed processes, or again concavo-convex, that side being 
convex which lies near the nuclear membrane (on a transverse 
section such a nucleus appears sickle shaped). The chromatin 
is arranged in the form of rather coarse granules (Figs. 339 
and 342), which after fixation in Hermann’s fluid (Fig. 340) 
appear to be the nodal points of a reticulum. 

The nuclei (Figs. 338-346) are numerous, vary in number 
from about six to about forty, and are irregular in size. Their 
shape is usually oval, seldom perfectly spherical, though quite 
frequently, as the figures show, they may be more or less 
elongate or even very irregular in form. Vacuoles are fre¬ 
quently present in them. The nucleoli stain as do all true 
nucleoli, but different degrees of staining density may be 
observed in the nucleoli of the same nucleus (Figs. 338, 342, 
346). In two cases, one of which is here figured (Fig. 342), a 
dense ring of chromatin was found around a nucleolus, but such 
cases, judging from their infrequency, must be regarded as very 
abnormal, if not attributable to the action of the fixing fluid. 
Divisions of the more elongate nucleoli certainly take place. 
Thus I have observed dumbbell-shaped nucleoli in three cases 
(Figs. 343, 345, 346), and Fig. 340 probably represents a stage 
just after a division, where two smaller nucleoli have apparently 
been divided off from a larger one, one end of the latter being 
drawn out to a point. Thus it might seem that the large num¬ 
ber of nucleoli are produced by divisions of a smaller number 
of larger nucleoli. The variability in the size, form, and 
number of these nucleoli recalls those of the subcuticular gland 
cells of Piscicola (cf infra) ; but in these cells of Doto I have 
been unable to make out different morphological phases. 

The cytoplasm of these cells is also remarkably differentiated 
(Fig. 339). In a given cell certain portions of the cytoplasm 


MONTGOMERY. 


[Vol. XV. 


482 

may be dense and stain deeply; other portions are less dense in 
structure, with a corresponding less intensity of stain; and still 
other portions of the cell substance appear structureless and 
do not stain at all. The cytoplasm in at least a portion of the 
peripheral area of the cell is always dense and deeply staining ; 
rarely is the cytoplasm in the whole cell of this dense structure. 
With low powers of magnification (e.g., Zeiss Obj. C, oc. 2 or 4) 
there may appear to be either several cavities in the cytoplasm 
or a single large one at one side of the nucleus. These differ¬ 
entiations of the cytoplasm (which fixation in corrosive subli¬ 
mate or in Hermann’s fluid bring out always in the same manner) 
probably denote certain metabolic states of the cytoplasm, but 
it would be difficult to determine from the structure alone to 
what physiological processes these states might correspond. 
There is no definite secretion produced by the cytoplasm, i.e ., 
no secretion with a definite form. As has been noted, a wholly 
or nearly wholly clear space often occurs in the cytoplasm at 
one side of the nucleus ; such a space usually lies at that 
margin of the nucleus situated closest to the center of the cell, 
and the nucleus may often surround it to some extent. Where 
the nucleus comes into contact with this space its membrane is 
thinnest and its outline irregular, and quite frequently this 
margin of the nucleus is produced into long, irregular, amoe¬ 
boid processes, which extend into the space in question and 
pass around it. These appearances would show that the 
nucleus stands in a certain functional relation to the metabolic 
changes of the cytoplasm, not improbably that it assimilates 
certain substances produced in the latter. 

To return to the nucleoli, I cannot find any genetic connec¬ 
tion between these structures and the cytoplasm. They are 
usually grouped near the center of the nucleus, and though 
often quite peripheral in position, never come into contact 
with the nuclear membrane, nor are they found in the amoeboid 
processes of the nucleus. It will be necessary to study very 
young individuals of this mollusc in order to determine the 
mode of nucleolar development. 

The cell (Fig. 339) is developed by a delicate membrane, 
which seems to be interrupted at no point on the surface of the 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 483 


cell. The cell has thus no external openings and no ducts or 
fibers which penetrate into the enveloping tissues. 

19. Subcutical Gland Cells of Piscicola rap ax (Verr.). 

(Plate 25 ; Plate 26, Figs. 198-203.) 

(These cells lie for the most part in the body cavity, between 
the body muscular wall and the intestine. Two modifications 
of them may be distinguished : (1) those at the ends of the 
body, near the suckers and in the wall of the latter, which are 
comparatively small, and the relatively short cell ducts of which 
open at all points of the surface of the body at the ends of the 
body and on the inner surface of the suckers; these seem to 
resemble the second modification in all respects except size ; (2) 
the larger type of these gland cells, which I have studied exclu¬ 
sively, are massed together in that portion of the body cavity 
which extends from the region a little anterior to the brain, 
nearly to the posterior end of the body, the greater number of 
them being in contact with the inner surface of the body 
wall.) 

In order to find all the functional stages of these cells one 
must study preparations of worms of various dimensions, since 
all the stages cannot be found in a single individual ; I made 
consecutive series of sections of seven different individuals, the 
smallest being about 6 mm. in length, and the largest being fully 
matured. The remarkable cycles of the nuclear and cell stages, 
to be described below, were equally well discernible with all 
three of the fixatives employed, namely, Flemming’s fluid and 
alcoholic and aqueous solutions of corrosive sublimate ; various 
double stains were used. 

These cells, when they reach their fullest dimensions, are so 
enormous that they may be readily seen with the naked eye. 
Their single ducts all open on the surface of the body, between 
the epithelial cells, a little anterior to the region of the sexual 
pore ; their openings are at this point equally numerous on 
the dorsal, lateral, and ventral sides of the worm. The most 
posterior gland cells of the body send their ducts a distance of 
four-fifths the total length of the body before they open on the 


484 


MONTGOMERY. 


[Vol. XV. 


surface of the latter, these ducts transversing a large number of 
body segments (in certain of the enchytraeid Oligochaeta there 
have been described subcutical gland cells whose ducts pass 
through a number of segments, but I believe that they are not 
of the same relative length as those of Piscicola). Each cell has 
its own duct, the latter being morphologically merely a process 
of the cell (Figs. 178, 181, 202); and as these individual ducts 
run in bundles parallel to one another, on their way to the sur¬ 
face of the body, they become closely apposed to one another, 
but there are apparently no open communications between the 
several ducts, nor do they unite to form larger, compound ducts. 
The ducts of those gland cells which are situated behind the 
sexual pore necessarily have an anterior direction, while those 
which are situated near to the head end of the animal send their 
ducts posteriorly. The duct departs from the cell more or less 
at right angles from its distal end, i.e., that end which is usually 
directed towards the central axis of the worm. Since the greater 
number of these cells become filled with secretion only when the 
worm is sexually mature, and since they all open on the surface 
of the body near the sexual pore, they have probably the same 
function as the clitellar glands of the Oligochaeta; after these 
observations had been completed I found that Bourne (’ 84 ) had 
described such gland cells in Pontobdella as “ clitellar glands,” 
but he made no observations on their finer structure. 

In studying the cycle of the structural changes of these 
cells two main morphological periods may be distinguished : 
(1) th z prophasis, from the immature cell to the cell filled with 
secretion ; and (2) the metaphasis , from the time when the 
cell begins to discharge its secretion until it becomes re-formed 
into a functionally immature cell again. I have no means of 
determining whether a given cell becomes filled with secretion 
only once a year (as, eg. y at the period of sexual maturity) or 
whether it may secrete several times in succession during the 
sexual period. At any rate, all appearances lead me to con¬ 
clude that it secretes periodically, most probably once during 
each period of sexual maturity. I have found no evidences that 
it secretes only once and then dies to become absorbed by 
the other tissues of the body; in other words, there were no 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 485 

evidences of cell degeneration or of a formation of new cells, so 
that we must conclude that each of these cells continues to 
functionate periodically during the whole time of the existence 
of the leech. 

We may now describe in succession the prophase and the 
metaphase of the structural changes. 

Prophasis (Figs. 178-196). — In the smallest cells found in 
the youngest leech examined no trace of secretion is present 
(Figs. 178-180). In these the nucleus is usually central in 
position, with a delicate chromatin network, and with a single, 
most frequently oval, nucleolus, in which one or a few small vac¬ 
uoles are commonly present. Around the nucleus, and filling 
the cell duct, is a somewhat dense cytoplasm, which becomes 
more vacuolar at the periphery of the cell. The chief cyto¬ 
plasmic changes from now on are as follows (I have not fig¬ 
ured these changes, since they may be briefly characterized) : 
that portion of the cytoplasm close to the nucleus gradually 
becomes more dense and begins to stain differently from the 
rest, and then becomes quite homogeneous ; most frequently 
there is a layer of this homogeneous substance between the 
nucleus and the cell duct, only that portion of the cytoplasm at 
the proximal end of the cell, as well as a thin layer around 
the homogeneous substance, retaining its primitive appearance. 
Next, this homogeneous mass gradually breaks up into the 
numerous secretion corpuscles (Fig. 181, Seer.), the shape of 
the latter being ovoid after fixation in corrosive sublimate, but 
more spherical after the action of Flemming’s fluid. These 
secretion corpuscles stain at first just like the homogeneous 
substance, but gradually commence to stain otherwise, and in 
the functionally mature cell stain differently from the primitive 
cytoplasm, as well as from the homogeneous substance from 
which they were derived. The whole cell thus gradually 
becomes filled with these small corpuscles, until finally no 
trace of the original cytoplasm is to be seen, except a few 
faintly staining fibers. The cytoplasm which fills the duct 
undergoes the same morphological changes as that of the cell 
body just described, so that the first secretion corpuscles in it 
are the derivatives of its own substance ; the cytoplasm of the 


486 


MONTGOMERY. 


[Vol. XV. 


duct and of the distal portion of the cell are as a rule the first 
portions to become differentiated into the secretion. At the 
end of the prophase the cell has attained its maximum size, 
and the duct its greatest diameter, both containing hundreds 
of the mature secretion corpuscles lying in an unstained, struc¬ 
tureless fluid. The duct in all stages is always larger at its proxi¬ 
mal than at the distal end, though it narrows very gradually. 

But the most interesting morphological changes are those of 
the nucleus. While the secretion is being produced in the 
cytoplasm the nucleus increases rapidly in size, and at the 
same time becomes very irregular in form, until in the nearly 
physiologically mature cell it attains enormous dimensions and 
sends out through the substance of the cell long branching 
processes, which anastomose with one another and some of 
which reach even to the cell membrane (Figs. 178-196). Kor- 
schelt has described (’ 89 ) branched nuclei in the spinning glands 
of certain insect larvae, which are somewhat similar to the 
nuclei here delineated. The nucleus attains its greatest dimen¬ 
sions and its most marked degree of ramification when there 
is the greatest amount of the homogeneous substance in the 
cell, i.e. y just before this substance becomes metamorphosed 
into the secretion corpuscles. At this stage we find the greater 
portion of the nucleus situated at the proximal part of the 
cell, and from that point it sends out irregular branches which 
envelop the mass of homogeneous substance, and which pene¬ 
trate into it. At this period, further, no two nuclei are alike 
in form, so that it would be in vain to attempt to figure all 
the shapes which they may assume. The nuclear membrane 
becomes very thin, often scarcely perceptible, around the 
branched processes. I know of no other nuclei which are more 
interesting in point of size and variability of form than these ; 
and it would well repay accurate investigation in the endeavor to 
decide in what way they may influence or modify the cytoplas¬ 
mic changes which are simultaneously taking place, for they 
obviously have a close physiological connection with the forma¬ 
tion of the cellular secretion. Since the nucleus undergoes a 
rapid process of growth in these stages, we are obliged to 
assume that it is taking up substances from the cell body ; but 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 487 

it probably does not assimilate the mature secretion corpuscles, 
since when the latter are produced, as we shall see, the nucleus 
commences to retract in size and to withdraw its processes. As 
the nucleus increases in size its chromatin reticulum becomes 
looser, as if it were elastically stretched by the expansion in 
volume of the nucleus; the chromatin is continued into the 
ramifying processes of the nucleus. 

The nucleolar changes during the prophase are as follows : 
in the immature cell there is invariably a single rather large 
nucleolus, which occupies a more or less central position in the 
nucleus (Figs. 178-181, 184) ; it may be either oval or spindle 
shaped, and most frequently contains one or several small vac¬ 
uoles. Its substance appears homogeneous after treatment with 
corrosive sublimate, granular after the action of the fluid of 
Flemming, and has no limiting membrane ; in all its stages 
within the nucleus it stains very intensely, though always dif¬ 
ferently from the chromatin. Now as the nucleus increases 
in volume so also does the nucleolus, though at first at a rela¬ 
tively more rapid rate than does the former ; and in growing 
larger it gradually becomes more elongated, rod shaped, and at 
this stage is most frequently in contact with the nuclear mem¬ 
brane (Fig. 182). When it has taken up this peripheral posi¬ 
tion its period of most rapid growth commences, so that at 
this stage there is a proportionately greater amount of nucleo¬ 
lar substance in the nucleus than at any other period in its 
history. When it is apposed to the nuclear membrane it has 
at first more or less the form of a rod (often of a slightly curved 
rod), but as its substance commences to increase in volume this 
rod shape gradually becomes changed and the nucleolus becomes 
bent inwards (towards the center of the nucleus), frequently in 
the form of a V, an S, or a W, though there is marked vari¬ 
ability in regard to the form it may assume, since no two nucle¬ 
oli can be found at this stage which have exactly the same form 
(Fig. 189). It is about this time that the nucleolus attains its 
greatest staining density. Then this large and irregularly 
shaped nucleolus leaves the nuclear membrane and begins to 
fragment into pieces, which are very irregular in shape and 
variable in number and size ; the nucleolus may show thereby 


488 


MONTGOMERY. 


[Vol. XV. 


a number of constrictions, or buds of nucleolar substance may 
project from its surface ; it may first break into two larger 
pieces, and then these may fragment further, or it may at once 
break into a number of pieces which are irregular in their 
dimensions (Figs. 185-188, 190, 191, 193). These fragments 
gradually wander apart from one another, the nucleus now 
being larger and already somewhat irregular in shape ; and at 
the same time each of the primitive nucleolar fragments divides 
into smaller pieces of unequal size, until when the nucleus has 
attained its greatest dimensions and most pronounced degree 
of ramification it contains a very large number of irregular 
nucleoli, which are unequal in their dimensions (Figs. 194- 
196). The figures given of this last stage show only sections 
of nuclei, and since as many as five or six sections may be made 
of one of these colossal nuclei (my sections were between 3 and 
5 /a in thickness), not one of these figures shows more than a por¬ 
tion of the total number of nucleoli in these largest nuclei ; in 
some of the latter nuclei I compute the number of the nucleolar 
fragments to be at least three hundred. But the total mass of 
nucleolar substance in these largest nuclei is certainly consid¬ 
erably greater than the mass of the primitive nucleolus at 
the time of its greatest size ; accordingly, though the division 
products of the primitive nucleolus might constitute the greater 
part of the nucleolar substance in the largest nuclei, they do 
not constitute all of it. Therefore there must be a formation 
of new nucleolar substance after the primitive nucleolus has 
divided, i.e ., a production of nucleolar substance not derived 
from the primitive nucleolus; I cannot determine the manner 
of formation of this new nucleolar substance, but would suggest 
that either new nucleoli are formed, or that the fragments of the 
primitive nucleolus increase in size by the addition of new nucle¬ 
olar substance to them. The greater number of nucleoli in the 
largest nuclei are collected in or near the thicker portion of the 
nucleus and few or none lie in the branched processes ; they are 
at this time seldom in contact with the nuclear membrane. Only 
a few of them contain vacuoles, and those which do may be re¬ 
garded as derivatives of the primitive nucleolus, the vacuoles of 
the latter still being preserved in its daughter-nucleoli. 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 489 

A nucleolar change now occurs which I have never seen 
paralleled, and to my knowledge no similar morphological change 
has ever been described. At the time when the homogeneous 
substance of the cell is commencing to differentiate into the 
secretion corpuscles, the nucleus begins to withdraw its branched 
processes and to decrease in size; while so doing it discharges 
its nucleoli into the cell body (Figs. 197-199). There can be no 
doubt of the genuineness of this process, since I have examined 
at least two hundred nuclei at this stage, which showed all 
intermediate stages between nuclei which had discharged only 
a few nucleoli and those which had discharged all except a 
single one of their nucleoli into the cell. The study of these 
nuclei gives the impression that successive contractions of the 
nucleus take place, whereby at first all the more peripheral nucle¬ 
oli, and later those which are more central in position, become 
successively extruded, for in the cell two or three more or less 
parallel rows of nucleoli may be found, or more properly speak¬ 
ing, concentric circles of nucleoli (Figs. 197 and 198). In some 
cases I have observed nucleoli which were halfway through 
the nuclear membrane, but by far the greater number of the 
nucleoli are found either within or without the nucleus, and 
this would prove that the contractions of the nucleus are sudden 
in their action. I think that it is the sudden contractions of 
the nucleus which alone cause the expulsion of the nucleoli, 
for as the nucleus diminishes in volume its chromatin network 
may be seen gradually to become closer and denser, and the 
pressure within the nucleus becoming greater than the pressure 
without it, the nucleoli, not being fixed in position, are forced 
out into the cell body where there is comparatively little pres¬ 
sure, since the secretion corpuscles are not densely grouped, 
but lie scattered through a thin and structureless fluid substance. 
The nucleoli, when they have arrived in the cell body, are 
not found in equal number at all points around the nucleus ; 
accordingly they are probably not discharged from all sides of 
the nucleolus in equal number, but only there where the nuclear 
membrane is thinnest (it is probably thinnest at those points 
whither the nuclear processes had withdrawn themselves). But 
though the nuclear membrane appears to be thinner at some 


490 


MONTGOMERY. 


[Vol. XV. 


points than at others, there are no visible pores in it, so that 
the nucleolar substance must be squeezed through the nuclear 
membrane itself. When one takes a sponge filled with water 
and presses it in the hand the water is forced out of it in the 
form of jets or columns, which are radial to the surface of the 
sponge; exactly similar seems to be the method of the discharge 
of the nucleoli in the case at issue, except that the nucleus is 
itself actively contracting. Thus we find the greater number 
of the nucleoli which lie in the cell body close to the surface 
of the nucleus to be irregularly columnar or rod like in shape 
(Fig. 198) and radially grouped around the nucleus. Those 
which lie nearer the periphery of the cell, however, and which 
had probably been discharged by a previous contraction of the 
nucleus, are more irregular in form, and their axes have a less 
regular position with regard to that of the nucleus. Further, 
those ends of the rod-like nucleoli in the cell which are directed 
towards the surface of the nucleus are usually more attenuated 
than the opposite ends, i.e., a nucleolus lying in the cell close 
to the nucleus has often the form of a pyramid the apex of 
which is directed towards the surface of the nucleus, and this 
form we would expect to result in the squeezing of a more or 
less viscid substance, like that of the nucleoli, through the 
nuclear membrane. I give only two figures showing the stage 
of the discharge of the nucleoli from the nucleus, simply in 
order to save time in the drawing of the numerous nucleoli ; 
but my preparations show very clearly all the stages of this 
process: one has only to examine sections of the mature leech to 
find them in abundance. The extrusion of the nucleoli continues 
until only about twenty, then a dozen, then four or five, and 
finally only a single nucleolus (Fig. 199) remains in the nucleus ; 
corresponding to these successive states of the discharge of the 
nucleoli we find cells in which only a few nucleoli, and then 
those in which the greater number of the nucleoli, lie in the 
cell body. One nucleolus always remains in the nucleus, though 
this one appears to differ in no wise from those which are dis¬ 
charged. Those nucleoli which lie in the cell body (Figs. 197- 
199) differ from those in the nucleus in their lesser density, 
greater size, and different reactions to certain stains (we shall 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 491 

return to the chemical change later); in other words, the sub¬ 
stance of those nucleoli which have come to be situated in the 
cell body undergoes a physical and perhaps a chemical change 
in this portion of the cell, and their expansion in volume might 
be accounted for on the ground of there being a smaller degree 
of pressure in the cell body than there is in the nucleus. 

It will be noticed that the prophase and the metaphase of 
the cell body and of the nucleus do not exactly coincide in 
point of time, the metaphase of the nucleus commencing earlier 
than that of the cell body. Thus the nucleus attains its great¬ 
est dimensions and most diverse ramification at the time when 
the cell body contains the greatest amount of homogeneous 
substance, and the nucleus enters on its metaphase (diminu¬ 
tion in volume, retraction of processes, expulsion of nucleoli) 
when the secretion corpuscles are only commencing to arise in 
the cell body. At the beginning of the metaphase of the cell 
body (when the latter is filled with the secretion corpuscles and 
commences to excrete them) the nucleus has already assumed 
a nearly spherical or oval form, has greatly decreased in size, 
and has discharged most of its nucleoli into the cell, i.e ., the 
nucleus has advanced already some distance in the path of 
the metaphasis. 

The metaphase of the cell body (Figs. 198-203) commences 
when the cell is filled with the secretion corpuscles, all traces 
of the previous homogeneous substance being absent, and begins 
to discharge them through its duct. During this process the 
cell gradually decreases in size, and the primitive cytoplasm 
again comes into view, at first in the form of delicate fibers. 
When the cell has shrunk to about one-third of its former size 
(the diameter of the duct does not decrease quite so rapidly, 
since it may be still full of secretion corpuscles after they have 
all disappeared from the cell body) the nucleus has simultane¬ 
ously decreased in size, but with greater proportionate rapidity 
than the cell body, and so at the close of the metaphase (Fig. 
202) the nucleus reaches its smallest relative size. The latter 
contains at this stage invariably a single nucleolus, of spherical 
or oval form, very regular in outline, and exactly similar to the 
nucleolus at the commencement of the anaphase-except that 


492 


MONTGOMERY. 


[Vol. XV. 


it does not appear to contain vacuoles. The nucleus itself is 
somewhat elongate and irregular in outline, and, owing to its 
maximum degree of contraction (a characteristic of the end of 
the metaphase), its chromatin builds a dense network within it. 
A study of the cell body at this stage allows us to follow 
the morphological changes undergone by those nucleoli which 
had been discharged by the nucleus (Figs. 198-203). The 
cytoplasm gradually assumes a reticulate or a somewhat granu¬ 
lar structure, and finally a most regular vacuolar or alveolar 
structure. As the cell body decreases in size the discharged 
nucleoli lying in it gradually stain less deeply, they lose their 
rod-like form, and no longer remain isolated, but all the nucleo¬ 
lar substance in the cytoplasm gradually becomes confluent, 
and becomes arranged in the form of a coarse, irregular network 
of substance distributed in the cytoplasm, and readily distin¬ 
guishable from the latter by its different staining properties 
(Figs. 201-203). By a hasty inspection this network of nucle¬ 
olar substance might appear to represent branches' of the 
nucleus, but a careful study shows that at this period of its 
growth the nucleus has no branches. As the cell continues to 
become smaller the amount of nucleolar substance in the cyto¬ 
plasm gradually becomes less and less, first the network at the 
periphery of the cell disappearing, then that in the vicinity of 
the nucleus, until at the conclusion of the metaphase no nucle¬ 
olar substance is any longer to be seen in the cytoplasm. I am 
unable to determine whether it is finally discharged through 
the cell membrane or whether it becomes metamorphosed into 
cytoplasm ; it certainly is not excreted through the cell duct, 
since no nuclear substance occurs in the latter, and at this 
stage the duct is no longer an open tube, but all the secretion 
corpuscles having been expelled from it, it is again filled with 
cytoplasm. The suggestion may be made that at least a portion 
of this nucleolar substance remains in the cytoplasm, so that in 
the succeeding prophase the nucleolus within the nucleus might 
find the material necessary for its growth in the nucleolar sub¬ 
stance suspended in the cytoplasm ; thus there might be, in the 
history of the nucleolar substance, periods of its expedition into 
the cytoplasm alternating with those when it is again taken 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


into the nucleus. And that in the prophase the single nucle¬ 
olus of the nucleus derives the material necessary for its 
further growth from the cell substance, seems highly probable 
when we recall the fact that at the time of its most rapid 
growth it is usually apposed to the nuclear membrane, which 
would denote that it is taking up a substance which penetrates 
that membrane from the side of the cell body. 

We have alluded to certain chemical changes which occur in 
the nucleolar substance when discharged from the nucleus 
during the metaphase of the latter. These staining differentia¬ 
tions and the coloration of the cytoplasm as observed on five 
different preparations are as follows (the first preparation was 
fixed with Flemming’s fluid, the others with corrosive subli¬ 
mate). 

First preparation (Ehrlich’s haematoxylin, two hours ; eosin, 
ten minutes): cytoplasm pale lilac; nucleoli in the nucleus, and 
when first discharged from it, reddish or rusty brown; nucleo¬ 
lar substance at the end of the metaphase lighter in color. 

Second preparation (gentian violet in aqueous solution, twenty- 
five minutes ; eosin, four and one-half minutes) : cytoplasm 
very faintly stained ; nucleoli in the nucleus deep violet, those 
in the cytoplasm yellowish red. 

Third preparation (Ehrlich’s haematoxylin, one hour ; eosin, 
five minutes): cytoplasm pale pink ; nucleoli in the nucleus, 
and when first discharged from it, purple ; nucleolar substance 
in the cytoplasm at the end of the metaphase pure blue. 

Fourth preparation (Ehrlich’s haematoxylin, forty minutes ; 
eosin, five minutes) : nucleolar substance within and without 
the nucleus yellowish red ; cytoplasm of a paler red. 

Fifth preparation (Mayer’s acid carmine, twenty minutes ; 
Lyons blue, five minutes): cytoplasm unstained ; nucleoli in the 
nucleus, and, when first discharged, bluish green; nucleolar 
substance at the end of the metaphase reddish purple in the 
cytoplasm. These methods of double staining show that the 
nucleolar substance, when discharged from the nucleus, under¬ 
goes some chemical change in the cytoplasm ; and they 
serve to distinguish, further, this substance from the true 
cytoplasm. 


494 


MONTGOMERY. 


[Vol. XV. 


I have no material of Piscicola after the breeding season, and 
accordingly could not follow the changes of these gland cells 
in their metamorphosis from the end of the metaphase to the 
commencement of the prophase. But these two end stages do 
not differ much fr^m one another, since the cell at the former 
stage differs from that of the latter merely in that its nucleus 
is smaller and more irregular in shape. 

It is not difficult to determine the sequence of the stages 
described ; only in the smallest individuals do all the stages of 
the prophase occur, and only in the largest those of the 
metaphase. 


20. Mesenchym Cells of Cerebratulus lacteus (Verr.). 

(Plate 29, Figs. 315a, 3^-324.) 

(I have described these cells in a previous contribution (’ 96 ), 
and so shall treat of them in this place mainly with regard to 
their nucleoli.) 

The smallest nuclei (Figs. 316a and 317) are densely filled 
with chromatin, and nucleoli appear to be absent ; the nuclear 
sap also stains with haematoxylin, so that these nuclei may 
be easily recognized by their deep stain and sometimes nearly 
homogeneous appearance. I have made a careful examination 
for nucleoli on preparations stained by the Ehrlich-Biondi 
method, as well as with haematoxylin and eosin, and am 
certain that nucleoli are either wholly absent or, if present, 
must be very minute in point of size. Such, then, is the 
structure of the smallest nuclei, namely, those found in the body 
cavity, and those of the smallest cells of the pseudoepithelia 
lining the body cavity. 

The non-continuous pseudoepithelia of the body cavity are 
layers of differentiated mesenchym cells, which differ from the 
primitive cells in their greater dimensions and more oval or 
spherical outlines (the undifferentiated cells are bipolar or 
multipolar, with long branching processes). In these larger 
cells we find for the first time a spherical, deeply staining 
nucleolus. Now the size of the latter stands in a pretty con- 
stent ratio to that of the nucleus. Further, in the smallest 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


nuclei which contain nucleoli, from one to three of the latter 
occur, and one or all of these are frequently found in close con¬ 
tact with the nuclear membrane (Figs. 315a, 318, 320), while in 
the largest nuclei observed only a single nucleolus is present, and 
this one is relatively large and is always at or near the center 
of the nucleus, never at its periphery (Figs. 319, 322, 323). 
In connection with the problem of the origin of this nucleolus 
we recall those small granules contained in the cytoplasm, 
which I have (’ 96 ) termed nutritive particles. These particles 
(Nut. 67 .) stain with eosin quite as intensely as the nucleolus, 
and in the smallest cells are either wholly absent or present in 
only small number ; but in the larger cells they are usually 
much more abundant, or when not more numerous they are of 
greater size, and are often quite densely grouped around the 
nucleus. It would seem probable that the nucleolar sub¬ 
stance is derived from these supposed nutritive particles. Thus 
when the nucleoli first appear they are most frequently in con¬ 
tact with the nuclear membrane ; and this shows that they are 
formed at the periphery of the nucleus, and only later come to 
occupy a central position within it. And since the nutritive 
particles are usually very numerous in the immediate vicinity 
of the nucleus, we may conclude that the nucleoli are formed 
from substance of these nutritive particles, which has been 
taken up by the nucleus. In the smallest nuclei alone do more 
than one nucleolus appear, so that the nutritive substance 
would seem to be taken into the nucleus from several points 
on its periphery, and then subsequently these several assimi¬ 
lated portions of nutritive substance may fuse together and so 
produce a single large nucleolus. Accordingly, the substance 
of the nucleolus would in this way appear to have an extra- 
nuclear origin. That these nutritive particles are being succes¬ 
sively absorbed by the nucleus is shown by the fact that the 
increase in the size of the nucleus and of the nucleolus go hand 
in hand. On the other side, these nutritive bodies in the cyto¬ 
plasm cannot be considered to be of nucleolar origin, since 
they usually make their first appearance in the cell body before 
a nucleolus arises in the nucleus ; and if they did have a nucle¬ 
olar origin, i.e ., if they were excreted portions of the nucleolus. 


496 


MONTGOMERY. 


[Vol. XV. 


we should expect to find the largest nucleoli in the smallest 
cells and the smallest ones in the largest cells. Further, the 
nucleolar substance cannot be regarded as a secretion of the 
nucleus itself, since this would leave unexplained the peripheral 
position which it at first occupies in the nucleus. Thus the 
mode of origin of the nucleolus in these cells would seem to be 
similar to that of the nucleoli in the ova of the nemerteans. 
A final point may be noted : the nucleolus accepts the same 
stains, though more intensely, than do the nutritive particles in 
the cytoplasm ; accordingly, the substance of those bodies which 
have been absorbed by the nucleus, and then by their fusion in 
the nucleus produce the nucleolus, must have undergone either 
a slight chemical or physical change within the nucleus. 

The largest mesenchym cells of the pseudoepithelia probably 
represent the youngest stages of the ova, though in the single 
individual of this species at my disposal no gonads were pres¬ 
ent, so that I can bring no proof positive that this is the mode 
of origin of the egg cells. In Carinella it is from similar cells 
that the genital products are derived, as I have previously 
shown (’ 96 ). Coe (’ 95 ) described certain of the more mature 
egg stages. 

In my earlier paper on these cells ( l.c .) I termed all the 
nuclear divisions of these cells “amitotic.” But renewed study 
of these elements shows that only the divisions of those cells 
are amitotic (Figs. 316a and 317), from which the free mesen¬ 
chym cells are produced. Whereas, in the nuclear divisions of 
the cells of the pseudoepithelia from which the masses of larger 
cells are derived I now find evidences of regularity in the 
distribution of the chromatin, so that probably these divisions 
are mitotic. However, in these small nuclear divisions it is 
almost impossible to decide whether we have to do with mitoses 
or with amitoses without the use of better lenses than those 
which were at my disposal. 

21. Ganglion Cells of Nemerteans. 

I may here briefly mention the relations of the nucleoli in 
these cells, and for other details refer to a previous contribution 
of mine (’ 97 ). 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


Linens gesserensis. —Cells of the first type : one or two small 
nucleoli. Cells of the second type : one nucleolus. Cells of 
the third type : a single nucleolus, or two of unequal size. 

Cerebratnlus lacteus. — Cells of the first type : as in the 
preceding species. Cells of the second type : one or two 
nucleoli. Cells of the third type : one or two nucleoli, which 
in one case stained differently. Cells of the fourth type : 
usually one peripheral nucleolus; rarely are two present, and 
then they are unequal in dimensions. 

In all these cells the nucleolus is comparatively small, homo¬ 
geneous, and no evidences of nucleolar division were seen. 

IV. GENERAL COMPARISONS AND CONCLUSIONS. 

Here I shall summarize merely the results of my observations 
on the nucleolus, and compare them with the conclusions of 
other investigators. Numerous other morphological points 
have been brought up, however, in the preceding pages, such 
as yolk development, differentiation of ova, nuclear divisions, 
distribution of the chromatin elements in the germinal vesicle 
at different stages in the growth period, changes in the struc¬ 
ture of cytoplasm, etc. 

i. Chemistry of the Nucleolus. 

I have made no special chemical study of these structures, 
except what may be learned from their reactions to stains. In 
the gregarines no substance could be demonstrated which chemi¬ 
cally corresponds to the chromatin of the metazoan cell ; 1 but the 
following table represents the mode of staining of true nucleoli 
in the somatic and germ cells of the Metazoa: 


Stain. 

Nucleolus. 

Chromatin. 

Del. or Ehrl. haematoxylin, eosin 

. red 

. blue. 

Ehrlich-Biondi stain . 

. maroon or red 

. green. 

Acid carmine, nigrosine 

. blue or greenish 

. red. 

Del. haematoxylin, cochineal 

. pink or red 

. blue. 

Safranin, gentian violet, orange . 

. yellow 

. blue. 

Schwarz (’ 87 ) distinguishes in 

plant cells pyrenin, the sub- 


stance of the true nucleoli, from the other nuclear substances 

1 That is, not to chromatin in the form of pure nucleic acid. 


498 


MONTGOMERY. 


[Vol. XV. 


and finds that it has a closer chemical affinity to the substance 
of the nuclear membrane (amphipyrenin) than to any other 
substance. Judging merely from the reactions of these two 
substances to stains I would agree in this point with Schwarz. 
Zacharias (’ 82 ) shows also for plant cells that the nucleolar 
substance is sui generis and is allied to plastin. O. Hertwig 
(’ 92 ) terms the nucleolar substance “ Paranuclein ” and observes: 
“ Nuclein und Paranuclein betrachte ich als die wesentlichen 
Substanzen des Kerns. . . . Beide scheinen mir in irgend 
welchen Beziehungen zu einander zu stehen.” But it is impor¬ 
tant to note that the true nucleolar substance probably has no 
chemical relation to the true chromatin (nuclein). Thus karyo- 
somes should not be considered as a particular group of 
nucleoli, since they are not nucleoli at all, but nodal points of 
the chromatin reticulum. The substance of every true meta¬ 
zoan nucleolus apparently differs chemically from the chromatin, 
linin, paralinin, and oedematin (lanthanin) ; and accordingly 
“pyrenin” is a term preferable to “paranuclein,” though 
“ pyrenin ” may include divers substances. 

There are also chemical differences between the nucleoli 
proper (“ Hauptnucleoli ”) and the paranucleoli (“ Nebennucle- 
oli ”), which occur together in many ova and yi a few somatic 
cells ; the substance of the paranucleoli stains more lightly 
than that of the nucleoli proper. List (’ 96 ) distinguishes three 
kinds of true nucleoli, from a chemical standpoint : (i) the 
nucleolus of somatic cells ; (2) the nucleolus proper of germinal 
vesicles ; and (3) the paranucleolus of germinal vesicles ; and 
he considers the substance of the paranucleus of the germ cell 
to be closer related chemically to the nucleolus of somatic cells 
than either of them is to the nucleolus proper of ova. List 
promises a more complete paper on this subject. The so-called 
“ nucleoli,” which react like chromatin, are of course not true 
nucleoli, but either karyosomes (thickened nodal points of the 
chromatin reticulum) or chromatin nucleoli (independent lumps 
or spheres of chromatin). It is my intention to devote a special 
paper to the consideration of the latter structures. Other 
papers on the chemistry : Macallum (’95), Michel (’ 96 ), Carnoy 
and Lebrun (’97). 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


499 


2. Number of Nucleoli. 

As Flemming (’ 82 ) has stated, the number of nucleoli is 
small in most cells, not more than from one to five. But in 
certain stages of some cells there may be several hundred (ova 
of Reptilia , Amphibia , Selachii , nemerteans, subcuticular gland 
cells of Piscicold). Even in those cases just mentioned, where the 
number of the nucleoli is very large, the immature cell contains 
only one or a few nucleoli, so that the large number is attained 
only when the nucleus has increased in size, cf. the observa¬ 
tions of Auerbach (’74a). Among somatic cells a large number 
of nucleoli is much more infrequent than among egg cells. At 
a given stage of a given cell of any one species of metazoan 
the number of nucleoli is pretty constant, and there is less 
variability in the number among those cells where the typical 
number of nucleoli is a small one than in those where a large 
number is present. In cells where the usual number of nucleoli 
is one or two, as in those of the nidamental gland of Mon - 
tagua , three may quite frequently be found, but no cells are 
found in which not a single nucleolus occurs ; in other words, 
there is in most cases some degree of variability in the number 
of the nucleoli, and the amount of this variability stands in a 
more or less direct ratio to the number of the nucleoli, but it is 
numerically progressive as a rule, tending to produce more than 
the normal number, and in no cases where cells normally con¬ 
tain nucleoli do we find a regressive numerical variation leading 
to the total disappearance of nucleoli. In certain few cells no 
nucleoli are present, and this is the case in more cells than 
Flemming (’ 82 ) was disposed to admit, since not only are spe¬ 
cialized cells like mammalian blood corpuscles without them, but 
they are also absent in certain connective-tissue elements of 
nemerteans, and in certain other cells of a low degree of vitality. 

Auerbach (’ 90 ) formulated the law that the number of nucle¬ 
oli is more or less constant for all the cells of a given species. 
But this conclusion is certainly erroneous, since in Doto there 
is one nucleolus found in the blood corpuscles and in the ovum, 
from one to five in the ganglion cells, from one to three in the 
cells of the nidamental gland, and in the giant cells as many as 


500 


MONTGOMERY. 


[Vol. XV. 


forty ; and in Piscicola , usually one in the ovum and the 
ganglion cells, about twelve in the mature muscle cells, and 
three hundred or four hundred in the subcuticular gland cells. 
From the data at hand we accordingly conclude that the 
number of nucleoli is not constant for the species. (On the 
number of nucleoli at different stages in amphibian ova, cf. Car- 
noy and Lebrun, ’ 97 a). 

In order to determine whether the number of nucleoli in egg 
cells were fixed for, or in any way determined by, the particular 
groups of Metazoa, I have compiled the following tables (pp. 501- 
505) for the larger groups, these tables representing the data 
of previous investigators and of my own observations. In them 
four classes of germinal vesicles are distinguished according to 
differences in the number and kind of the nucleoli; this classifi¬ 
cation is only for convenience’ sake, only arbitrarily chosen, and 
is probably not a natural one. On the left hand is given the 
name of the genus or group ; the asterisk corresponding to each 
form indicates by its position in a particular vertical column the 
nucleolar relations of the ovum of the form specified ; and next 
to the asterisk is placed the name of the authority. In some 
cases two investigators may have reached different conclusions 
in regard to the nucleolar relations, so that for these cases two 
asterisks were employed. 

One must be extremely cautious in any attempt to draw 
conclusions from these data, not only because the data are 
so meager, but also because where data have been culled 
from so many different observers some of the facts may ulti¬ 
mately prove to have been erroneous. Thus many of these 
ova may have been examined at only one point in their develop¬ 
ment, and in others paranucleoli may have been entirely over¬ 
looked, or may have been confused with true nucleoli. But 
taking this mass of observations as it stands, the following gen¬ 
eral conclusions may be drawn: we find that a large number of 
nucleoli is not always characteristic of ova with a considerable 
amount of deutoplasmic substances, for a single nucleolus is 
typical for the birds and for many of the Arthropoda. Further, 
the number of the nucleoli does not seem to be dependent 
upon the amount of yolk, nor upon the mode of cleavage, 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 501 


Form. 

A Single 

More than One Nu¬ 

Nucleolus and 

N UCLEOLUS. 

cleolus, All Alike. 

Paranucleolus. 


Coelenterata . 


f Esperella 

* (H. V. Wilson) 



J Spongilla 

1 Tedamione 

* (Fiedler) 

* 1 


[_ Hircinia 
’ Hydrozoa 

* (Weismann) 

# \ (H.V. Wilson) 


Aequorea 

* (Hacker) 



Hydractinia 

Podocoryne 

# | (Bunting) 



- Geryonia 

* (Fol) 



Tubularia 

Eucope 

Aeginopsis 

* (Doflein) 

* (0. Hertwig) 

* (O. Hertwig) 

k Hydra 



* (Brauer) 

( Nausithoe 



*1 

\ Pelagia 



* l (O. Hertwig) 

J Physophora 
\ Rodalia 



*J 

Ctenophora 

* (Chun) 


* (mihi) 


Plathelminthes. 


Bothriocephalus 

Distomum 

* (Schauinsland) 

* (Schauinsland) 

Polycladidea 

* (Lang) 


Tricladidea 

Rhabdocoele 

* (Repiachoff) 

* (Jijima) 

Prorhynchus 

Bothrioplana 

# j- (Vejdovsky) 


Haplodiscus 

* (Bohmig) 



Nemertini. 


Carinella 

* (Burger) 




( Cerebratulus 

* (Hubrecht) 



* (Burger, Coe) 

\ Lineus 

* (mihi) 




r Malacobdella 


*< 

v. Kennel) 


Drepanophorus 

* (Burger) 




Tetrastemma 


*' 



Amphiporus 


* 



4 Stichostemma 


* 

► (mihi) 


Zygonemertes 


* 



Proneurotes 



l 


Prosadenoporus 


* (Burger) 


„ Pelagonemertes 



t 

* (Hubrecht) 
























502 


MONTGOMERY. 


[Vol. XV. 


Form. 

A Single 

More than One Nu¬ 

Nucleolus and 

Nucleolus. 

cleolus, All Alike. 

Paranucleolus. 


Annelida. 


' Nereis 



* (E. B. Wilson) 

Spinther 

Ophryotrocha 

% j- (Korschelt) 



Sternaspis 

Polydora 

* (Vejdovsky) 


* (mihi) 

Spio 



* (Giard) 

Capitellids 



* (Eisig) 

^ Polygordius 



* (Fraipont) 

( Oligochaeta 
■s Rhynchelmis 

L Lumbricus 

* 1 

# > (Vejdovsky) 


* (Clapar£de) 

' Nephelis 

* (Leydig) 



Branchiobdella 

* 1 



Haemopis 

* | (Ludwig) 


* (O. Hertwig) 

Piscicola 

Clepsine 

* (mihi) 

* (Whitman) 


w Piscicola 


* (Leydig) 



Arthropoda. 


' Homarus 

* (Herrick) 



Porcellio 

* (St. George) 



Oniscus 

* (Wielowiejski) 



Heterocope 

Diaptomus 

Argulus 

# j* (Riickert) 

* (Leydig) 


Astacus 


* (Wielowiejski) 


Cyclops 




Sida 

Canthocamptus 



* l (Hacker) 

Moina 



*J 

Euchaeta 



* (vom Rath) 

' Epeira 

* (Korschelt) 



Dolomedes 

* (Korschelt) 



Phalangium 

* (Korschelt, 




Leydig) 


* (Henking) 

Lycosa 

* (Leydig) 



Theridium 

* (v. Wittich) 


* (Leydig) 

Tetragnatha 



* (Leydig) 

A raneina 



* 1 

A carina 



# > (Wielowiejski) 

v Zilla 



* (Van Bambeke) 




















No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


503 


Form. 


A Single 
Nucleolus. 


More than One Nu- Nucleolus and 
cleolus, All Alike. Paranucleolus. 


' Julus 
Geophilus 
Glomeris 
^ Lithobius 
Peripatus 
' Blatta 
Nepa 
Notonecta 
Carabus nemoralis 
Gryllotalpa 
Pieris 
Anabolia 
Bombus 
Anomalia 
Ophion 
Ephialtes 
Pemphigus 
- Musca 

Necrophorus 
Geotrupes 
Banchus 
Pimpla 
Stenobothrus 
Meloe 
Libella 
Melolontha 
Lina 
Lycus 
Sphinx 
_ Ambyteles 


* (Stuhlmann) 


* 

* 

* 

* 

* 

* 

* 

* 

* 

* 

* 

* 


(Stuhlmann) 

(Brandt) 

} (Will) 


■ (Stuhlmann) 


* (Leydig) 

* (Wielowiejski) 


* (Balbiani) 


:i 

^ y (Stuhlmann) 

* J 

1 1 (Leydig) 


* (Leydig) 

* (Stuhlmann) 

* (Leydig) 


* (Stuhlmann) 


* 

MU 

MU 

* 

* 

MU 


(St. George) 
(Wagner) 

► (Stuhlmann) 


Psammechinus 

Echinocardium 

Echinus 

Toxopneustes 

Asteracanthion 

Sphaerechinus 

Amphidetus 

Solaster 


Echinodermata. 
* | (Bergh) 


* | (Ludwig) 


# j- (Hacker) 

* } (O. Hertwig) 


Mollusca. 


r Chaetoderma 

* (Wiren) 



\ Proneomenia 



* (Hubrecht) 




























504 


MONTGOMERY. 


[Vol. XV. 


Form. 


A Single 
Nucleolus. 


More than One Nu- Nucleolus and 
cleolus, All Alike. Paranucleolus. 


' Neritina 
Doto 
Montagua 
Tellina 
Helix 
Limax 
Arion 
Doris 
Aeolidia 
Amphorina 
„ Paludina 
" Anodonta 
Unio 
Mytilus 
Pholas 
k Cyclas 


* (Blochmann) 

* (mihi) 

* (Platner) 


* (mihi) 

* | (O. Hertwig) 

* (Mark) 

* (Platner) 

* J- (Lonnberg) 

* (Trinchese) 
(Leydig) 

* (Flemming) 

* (Hessling) 

* (Lonnberg) 

* (List) 

* (Stepanoff) 


Tunicata. 


Distaplia 

* (Davidoff) 


Phallusia 

* (Bergh) 


Botryllus 

* (Pizon) 


Clavelina 

* (Seeliger) 


Ciona 



Styelopsis 



Ascidia 




Vertebrata. 


Amphioxus 

* (Van d. Stricht) 


Petromyzon 

* (Bohm) 


f Scyllium 



-1 Torpedo 


* y (Ruckert) 

[ Pristiurus 


*J 

' Scorpaena 


* (Van Bambeke) 

Conger 


*i 

Gadus 


* l (Scharff) 

- Trigla 


*J 

Gasterosteus 


* (Ransom) 

Anguilla 


* (Brock) 

Cyprinus 


* (Eimer) 

f Rana 


* (Born) 

-j Amblystoma 


* (Fick) 

[ Triton 


* (Leydig) 

r Emys 


* (mihi) 

■j Lacerta 


* (Eimer) 

[ “ Turtle ” 


* (Agassiz) 


* 

* 

* 


(Floderus) 
(O. Hertwig) 























No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


Form. 


A Single More than One Nu- Nucleolus and 

Nucleolus. cleolus, All Alike. Paranucleolus. 


{ 


Gallus 

Fringilla 

Columba 

Felis 

Cavia 

Mus 

Vespertilio 

Sus 

Myoxus 

Talpa 

Ovis 

Lepus 

Homo 


* (Holl) 


* (v. Wittich) 

* (mihi) 

* (St. George) 


* (Rein) 

* (Holl) 


* (v. Beneden) 



* V (Leydig) 


* (Nagel) 


* (St. George) 

* (Flemming) 


nor yet upon the mode of deposition of the egg ( i.e ., whether 
it is pelagic, hatched in a cocoon, or nourished in an uterus). 
These facts hardly warrant an attempt to explain the factors 
limiting the number of nucleoli, and perhaps such explanations 
should rather be expected from experimental workers than from 
purely structural observers. On examining the metazoan groups 
in detail we find in certain of them a degree of uniformity in 
regard to the number of nucleoli. Thus the only vertebrate 
ova with two kinds of nucleoli are those of Lepus and Ovis. A 
single nucleolus is the rule for Amphioxus , Petromyzon , the 
birds, and most of the mammals ; the Reptilia , Amphibia , 
Teleostii, and the Selachii have numerous nucleoli. In the 
Tunicata there is either a single nucleolus or a nucleolus and 
paranucleoli; this is also the rule for the Echinodermata , 
Mollusca, and Annelida. In the Arthropoda there is consider¬ 
able diversity in regard to the number and differentiation of the 
nucleoli. In the nemerteans we find most usually either a 
single nucleolus or a large number of small ones. In the 
Plathelminthes one or two is the rule ; this is also most 
frequently the case for the coelenterates, but in some of the 
latter paranucleoli have been described. 












506 


MONTGOMERY. 


[Vol. XV. 


3. Position of the Nucleolus in the Nucleus. 

Where a single nucleolus is present it almost always lies 
excentrically, though not against the nuclear membrane. Those 
cases where it regularly occupies the center of the nucleus must 
be regarded as exceptional ; thus I am unable to agree with 
Macfarlane that the nucleolus is either the morphological or 
the tropic center of the cell. At the time of its origin, and 
often at the time of mitosis, the nucleolus may be in contact 
with the nuclear membrane. Where a number of nucleoli are 
present they may be scattered irregularly through the nucleus, 
or grouped at one point in it, or be concentrically arranged ; 
their position is often dependent upon the stage of the develop¬ 
ment of the nucleus. Thus in the metanemerteans examined 
by me they lie at the periphery in the smallest germinal 
vesicles, then wander towards its center, and finally migrate to 
the periphery again. 1 

The nucleoli lie in the nuclear sap, as a rule not in any close 
connection with the chromatin reticulum. But in those cases 
where the nucleolus may be unusually large it appears to be 
suspended by the fibers of this reticulum, but not in such a way 
that the fibers penetrate into its substance, but become simply 
wound around its surface ; thus it appears that when the 
nucleolus increases in size it forces apart the fibers of the 
nuclear network in such a way that the latter gradually pro¬ 
duce a latticework on its surface. In this way the nucleoli 
may be more or less held in position in the nucleus, but 
Herrick’s observations on the gravitation of the nucleolus 
show that it is not firmly held by the chromatin fibers. The 
nucleolus is, as it were, a ball lodged in the branches of a 
tree, its movements hindered by the intervening branches, but 
nevertheless not immovable. Various views on the mode of 
suspension of the nucleolus : Pfliicke (’95), Heidenhain (’92), 
Rosen (’95), Jensen (’83), Zimmermann (’96). Note also its 
peculiar position in Synapta (Leydig, ’52). 

1 For the opinions of other authors, cf. the reviews of the papers of Pfliicke 
(’95), Heidenhain (’92), Rosen (’95), Jensen (’83), Leydig (’52), Zimmermann 
(96), Schneider (’91). 


No. 2.] 


COMPARATIVE CYTOLOGICAL STUDIES. 


5 ° 7 


4. General Morphological Structure of the Nucleolus. 

The ground substance of the nucleolus is more or less dense, 
but not brittle, and either homogeneous or finely granular, 
rarely coarsely granular. It may be either * fluid or viscid in 
consistency. 

In the greater number of cases it has no limiting membrane. 
Such a membrane was found by me only in the germinal spot 
of Polydora , and here it appeared to be merely a denser portion 
of the ground substance. When any small nucleolus is viewed 
in its totality a membrane appears to surround it, but this 
phenomenon is due to the refraction of light from its convex 
surface, and many observers have been misled by this appear¬ 
ance into supposing that a membrane is present. Others in 
describing those states of nucleoli in which a large vacuole is 
present have erroneously described the peripheral layer of true 
ground substance as a nucleolar membrane ; it is necessary to 
distinguish between such a peripheral layer, which consists of 
true ground substance, and a nucleolar membrane proper, which 
is a differentiation of the ground substance. Some authors, 
eg., Lavdowsky (’94), have described a membrane of chromatin 
enveloping the nucleolus, and I have found that those of the 
giant cells of Doto may sometimes be surrounded by a mass of 
chromatin. But this apposition of a mass of chromatin in Doto 
is certainly an artefact, though it would seem probable that 
the nucleolus in some cases has an envelope of chromatin 
forming a distinct capsule separated from the chromatin net¬ 
work of the nucleus. I am able, however, to corroborate the 
observations of Macfarlane (’8i) and Pennington (’ 97 ), that the 
nucleolus in Spirogyra has a true membrane. 1 

A very unusual structure of the nucleolus is that afforded by 
the salivary gland cells of Chironomus as described by Balbiani 
(’8i), Leydig (’83), Korschelt (’84), and Macallum (’ 95 ). C. 
Schneider (’ 91 ) supposes the nucleoli, as well as the rest of the 
nuclear substance, to consist of “ Geriist ” (linin ?) and chro- 

1 The following writers have described nucleolar membranes: Macallum (’95), 
Carnoy and Lebrun (’97a), Will (’85), Holl (’93), Roule (’83), Burger (’90), 
Ogata (’83), Vejdovsky (’82), Meunier (’86), Carnoy (’86), Mann (’92). 


5°8 


MONTGOMERY. 


[Vol. XV 


matin, and considers the nucleoli to be only isolated masses of 
chromatin surrounded by linin sheaths ; these observations 
have not been corroborated by any other writers and would seem 
to be due to faulty methods of fixation. 

In opposition to Meunier (’86), and in agreement with most 
investigators, I must conclude that vacuoles are normal struc¬ 
tures in nucleoli, since they may be seen after the most diverse 
methods of fixation, and their size and number are not only to 
some extent limited for the particular cell, but are also different 
at different periods in the metamorphoses of the nucleus. It is 
the rule that the youngest nucleoli are homogeneous, and that 
vacuoles first arise when they have increased in size. Their 
size and number vary at different phases in the development 
of the nucleolus. Very frequently a number of smaller ones 
appear, and then these subsequently fuse together and produce 
a larger one. The nucleoli of egg cells are characterized as a 
rule by more numerous or larger vacuoles than those of somatic 
cells, and in many somatic cells these vacuoles appear to be 
wholly absent. The vacuolar substance appears in some cases 
not to be a derivative of the ground substance of the nucleolus, 
but to be derived from without the nucleolus (ova of Doto and 
Montagua). Perhaps this vacuolar substance always has an 
extranuclear origin, since in many cases a germinal spot grows 
larger merely by an increase in its volume, while the ground 
substance seems neither to increase nor diminish. 

The alveolar structure of nuclei as described by Purcell (’94), 
Schaudinn (’94), Korschelt (’ 95 ), and Lauterborn (’95b) is prob¬ 
ably referable to the regular distribution of equal-sized vacuoles 
in the nucleolus. 

A “ Kernkorperchenkreis,” a shell of minute granules 
arranged concentrically around the nucleolus, has been de¬ 
scribed by Eimer (’71, ’72), Auerbach (’74a, who considered it 
to be the result of opposing repulsive forces of the nucleolus 
and nuclear membrane), Brass (’89), Pfliicke (’ 95 ), Platner (’89a), 
Smirnow (’ 90 ), Engelmann (’80), Carnoy and Lebrun (’97a). A 
more or less similar phenomenon has been described by me for 
ganglion cells of Doto. Such a nucleolar circlet must be con¬ 
sidered, in most cases at least, an artefact. But in this cate- 


No. 2 .] COMPARATIVE CYTOLOGICAL STUDIES. 509 

gory should not be classed small masses of nucleolar substance 
grouped around a larger one, these being normal phenomena 
during the growth period of nucleoli. 

Reticulations within the nucleolar substance have been de¬ 
scribed by some few authors. Thus Carnoy (’85, ’97a), Meunier 
(’86), and Moll (’ 93 ) described nucleoli containing a skein of 
chromatin, but Zacharias and Strasburger (’88) did not find 
anything resembling these supposed skeins described for 
Spirogyra. Leydig (’88) states that the germinal spot of Lycosa 
“ bietet das Bild eines Knauels dar.” Fromann (’84) described 
the nucleolar substance as consisting of granules connected by 
fibers, Butschli (’80) found the nucleoli of Dinoflagellata to 
contain a fine reticulum, and Davidoff (’89) states that the 
germinal spot of Distaplia takes up portions of the nuclear 
reticulum into itself (but cf. Bancroft, ’98, and Shafer, ’80). The 
only structure which was found by me to resemble a skein was 
present in the later stages of the germinal spot of Polydora ; 
but in this object, owing to the gradual confluence of the vacu¬ 
oles, which thus produce anastomosing channels of vacuolar 
substance in the ground substance of the nucleolus, it is the 
true ground substance which represents a skein-like appear¬ 
ance. It is very probable that Carnoy and his followers have 
mistaken the vacuolar substance for the ground substance, and 
have considered the true ground substance to be chromatin ; 
I am forced to conclude that in all probability there are no 
skeins of chromatin lying in any metazoan nucleolus, since I 
have never found any evidence of chromatin in it in any meta¬ 
zoan cell. But it is not improbable that in the nuclei of 
gregarines chromatin may be massed in some or all of the 
nucleoli. 

Nucleolini, granules within the nucleolus, have frequently 
been observed. A single nucleolinus to a nucleolus has been 
described by Vejdovsky (’95b), Morgan (’96), Agassiz (’ 57 ), 
Kleinenberg (’ 72 ), Leydig (’88), Macfarlane (’85), Lavdowsky 
(-98), A. Brandt (’78), Van Bambeke (’86), Kosinski (’87, 33 ) ; 
several nucleolini to a nucleolus, by Burger (’ 90 ), Rhumbler 
(’93), Holl (’93), Wolters (’ 91 ), Schron (’65), Scharff (’ss), See- 
liger (’82), Gjurasin (’93), Haeckel (’ 74 ), Mann (’ 92 ), Van Bam- 


MONTGOMERY. 


[Vol. XV. 


510 

beke (’97b), Mark (’ 77 ), Bancroft (’98). Compare also the follow¬ 
ing : Huie (’ 97 ), Van Bambeke (’ 97 ), Kosinski (’87, ’93), Mark 
(’77), Zimmermann (’96), Hodge (’94). I have found these bodies 
occurring in varying number, though most frequently absent, 
in the nucleoli of various cells, and they appeared to be merely 
loosened portions of the ground substance which had come to 
lie within a vacuole. Macfarlane and his pupil Mann have 
described nucleolini under the names “ endonucleolus ” and 
“ nucleolo-nucleus ” as occurring singly and with great con¬ 
stancy in certain plant cells, though Zacharias (’85) studied 
Macfarlane’s object ( Chara ) and makes no mention of any of 
these structures. Macfarlane ascribes the utmost importance 
to his “endonucleolus,” regarding it as the tropic center of the 
cell and as an important mechanical agent during nuclear 
division. Mann has not only described a most complex struc¬ 
ture of the nucleolus, such as no other observer has yet seen, 
but also has found fine fibrils radiating out from it, which he 
supposes to penetrate through the nuclear cavity. From my 
own observations, and in agreement with the majority of ob¬ 
servers, I can attach no particular morphological significance to 
the nucleolinus; it appears to be only a detached portion of 
the nucleolar ground substance, to be in most cases absent, 
and when present to vary greatly in regard to size, position, and 
number. It is undoubtedly the case that many structures 
which have been described as nucleolini are in reality minute 
vacuoles, which from their refrangibility appear to be granules; 
such is the case with the minute vacuoles of Polydora and 
Montagna when studied after the action of certain stains, and 
has been shown for other objects by Zimmermann and Huie, 
Lavdowsky found in the nucleolus a central vacuole, and in 
the latter a small granule, which he supposed to be “das noch 
in Entwicklung begriffene Centrosoma,” destined to finally 
pass out of the nucleolus; he was unable to determine how 
it does wander out of the nucleolus and become the centrosome, 
so that his suggestion has merely the value of a hypothesis. 
Van Bambeke describes the nucleolinus of the germinal spot 
of Amaurobins as “ doue d’un mouvement tres vif ” ; this 
interesting phenomenon certainly deserves investigation, though 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 511 

it is not impossible that the supposed nucleolinus was in reality 
a microorganism inclosed in the vacuole of the nucleolus. 
(Cf. also Flemming’s observation on the egg of Ascidia , ’ 97 .) 
Supposed nerve fibrils in the nucleolus have been described by 
Eimer (’ 73 , ’ 90 ). 


5. Polarity of the Nucleolus. 

In the gregarine (Gonospora f) from the intestine of Lineus 
gesserensis it is the rule that the vacuoles make their first 
appearance at that pole of the nucleolus which is nearest to the 
nuclear membrane. In the germinal spot of Montagna the 
opposite position of the large excentric vacuole is the rule, 
though the percentage of cases in which the vacuole has a 
particular position with regard to the nuclear membrane is 
less than in the gregarine. On the contrary, in the germinal 
spots of Piscicola and Rodalia there is no regularity in regard 
to the position of the vacuoles, and in that of Polydora the 
vacuoles are, at the time of their first appearance, usually 
central in position. In the germinal spots of many other 
Metazoa , where a single large vacuole is present it more 
usually lies excentrically than centrally, though its position 
appears to be independent of the proximity of the nuclear 
membrane ; so that in these cases we can speak of a certain 
polarity in regard to the position of the vacuole within the 
nucleolus, and not of a polarity of the axis of the nucleolus 
in regard to the position of the nuclear membrane. But 
in the two gregarines examined by me the substance of the 
nucleolus, or of some of the nucleoli, is differentiated at 
two poles of the nucleolus, so that the portion of the ground 
substance at one end stains differently from that of the other 
end of the nucleolus ; this state apparently does not occur in 
the nucleoli of metazoan cells. It remains to be solved whether 
in the gregarines the chromatin or its physiological equivalent 
is localized at some particular point or pole of the nucleolus, 
i.e., whether or not such nucleoli should be compared to the 
nucleoli of the Metazoa. 


S 1 * 


MONTGOMERY. 


[Vol. XV. 


6 . Amoeboid Movements, Divisions, Fusions of Nucleoli. 

Amoeboid movements have been seen in life in metazoan 
cells by the following observers (germinal vesicles): A. Brandt 
(’74, Blatta), Eimer (’75, Silurus), O. Hertwig (’76, Rana, 
Pterotrachea ), La Valette St. George (’66, Libella; ’83, Isopoda ), 
Bergh (’79, Gonothryaea), Van Beneden (’69, ’76, Polystomum, 
Rana), Balbiani (’64, several genera of spiders), Leydig (’83, 
Libella), A. Brandt (’78, numerous Insecta, Distomum) f Van 
Bambeke (’86, Blatta), Knappe (’86, Bufo), Auerbach (’74a, 
Teleostii). In somatic cells : Schwalbe (’76, sympathetic gan¬ 
glion cells of Rana), Kidd (’75, epithelial cells from the mouth 
of Rana), Hodge (’94, nerve cells of Rana), Auerbach (’74b, 
salivary gland cells of Musca). In Protozoa: Van Beneden 
(’69, ’76, Gregarina, Monocystis). In plants, Zacharias (’85) has 
observed amoeboid movements in the nucleoli of Chara (an 
observation overlooked by Zimmermann, who states that such 
movements have not been seen in plants). 

These observations would show that amoeboid movements 
are probably natural phenomena of certain nucleoli, but one 
should not be too positive of the naturalness of these phe¬ 
nomena, since some of the observations were made upon the 
heated stage, and in all of them the object was probably more 
or less compressed and placed in artificial conditions. But they 
are in all probability frequently normal phenomena, since, as we 
shall see, divisions and fusions of nucleoli are certainly normal 
and of wide occurrence, and the latter can only be classed as 
forms of amoeboid motion. The question arises, Are these 
movements wholly passive, caused by movements in the other 
parts of the nucleus, or should they be considered an inherent 
function of the nucleolus ? The latter alternative would seem 
the more probable, since no movements of the other nuclear 
elements are known in the resting cell. Van Beneden (’69) has 
described rhythmic expansion and contraction of the volume of 
nucleoli in gregarines. But all these movements of nucleoli 
should not be regarded as automatic motions of the nucleolus 
in the sense that an Amoeba forms and retracts processes ; but 
rather with Rhumbler (’ 93 ) they should be regarded as “ Auf- 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 513 

losungsvorgange,” due to chemical changes in its substance. 
Cf. the movements described by Flemming (’ 97 ) for the ovum 
of Ascidia . 

The nucleolus has in some cases a viscid consistency (as 
described by me for Stichostemma ) and then may be irregular 
in form ; in other cases it is more fluid, and this is probably 
the case when it has regularly a spherical shape, i.e., the globu¬ 
lar form characteristic of drops of a thin liquid. Its more or 
less fluid consistency allows changes of form, division into 
particles, and fusions of neighboring nucleoli. 

The division of a nucleolus into two or more parts is a 
normal and regular phenomenon in many cells, though all 
nucleoli do not show this property. Two kinds of nucleolar 
division may be distinguished : ( 1 ) that mode by which the 
nucleolus becomes elongated and then breaks into two or more 
parts, whereby the daughter-nucleoli are usually capable of 
further division ; and ( 2 ) that mode by which the nucleolus 
fragments nearly simultaneously into a number of small gran¬ 
ules. From my own observations the former mode is evinced 
by the nucleoli of the muscle and giant gland cells of Piscicola , 
the giant cells of Doto , and the germinal spots at certain 
stages in the ovogenesis of the metanemerteans. This mode of 
division cannot be regarded as a phenomenon of nucleolar 
degeneration, since the nucleolus and its products may often 
continue to increase in size during the process of division. But 
the second mode, that by which the nucleolus breaks into a 
large number of granules, since it is particularly characteristic 
of the nucleolus in nuclear division, may be regarded as a 
process of degeneration ; the case of divisions during nuclear 
division shall be considered later. A strange mode of nucleolar 
division has been described by A. Schneider (’83). According 
to his observations on Klossia , the smaller nucleoli are portions 
of the inner substance of the larger nucleoli and wander out of 
each larger one by passing through the pore (“ canal micropy- 
laire ”) of the cortical substance of the latter; this intranucleolar 
origin of the smaller nucleoli is still open to question, since it 
was not observed in life, and since the canal micropylaire was 
observed in only one nucleolus. Marshall (’92) has described 


MONTGOMERY. 


[Vol. XV. 


5H 

a somewhat similar method of formation of the smaller nucleoli 
of Gregarina blattuvum. Now I found in the nucleus of the 
gregarine from Lmcus numerous nucleoli of different dimen¬ 
sions, and often irregular in their outlines ; and this irregularity 
in form would point not only to amoeboid movements of the 
nucleoli, but also to nucleolar divisions, since in the largest 
nuclei we find a large number of small nucleoli. All appear¬ 
ances showed that these smaller nucleoli are division products 
of the larger ones ; but it seems that they simply bud off from 
the surface of the latter, and are not preformed in their interior. 
In other words, Schneider and Marshall are probably correct 
in concluding that the smaller nucleoli are disassociated portions 
of the larger ones ; but they may perhaps be mistaken in 
assuming that they are preformed in the interior of the latter, 
since these investigators may have mistaken vacuoles for intra- 
nucleolar nucleoli. (Other observations on nucleolar divisions 
in resting cells : Hermann, ’89 ; Vejdovsky, ’95a ; Biitschli, ’80; 
R. Hertwig, ’76 ; Kultschitzky, ’88 ; Bergh, ’79 ; Bannwarth, ’92 ; 
Stuhlmann, ’86 ; A. Brandt, ’78 ; Scharff, ’88 ; Eisig, ’87 ; 
Cunningham, ’95 ; Kosinski, ’87, ’93 ; Carnoy and Lebrun, ’97a ; 
Steinhaus, ’88 ; Cuenot, ’91 ; Metzner, ’94.) 

Fusions of nucleoli are not as widely known as divisions, but 
there are some facts which would show that the former processes 
are by no means unusual in their occurrence. Such fusions 
have been described for cells of plants by Zacharias (’85), Mann 
(’92), and Wager (’93) ; for animal cells by Rhumbler (’93, ’95), 
Brauer (’9l), Leydig (’ 50 ), Pfitzner (’83), and Riickert (’92). I 
have found fusions of the nucleoli to be characteristic phenomena 
of certain stages in the maturation of the germinal vesicles of 
nemerteans, an extreme case being furnished by Stichostemma , 
where sometimes all the nucleoli may fuse together at the 
center of the nucleus, and so produce a single large one. The 
nucleolus at the time of its origin may be said to be undergoing 
a process of fusion, since it is produced by the coalescence of 
numerous smaller portions of nucleolar substance. There is 
nothing problematical in regard to the fusion of nucleoli, since 
it is a physical property for bodies of like nature (when fluid) 
to fuse together when they come into contact, though this 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 515 

process is to some extent dependent upon the nature of the 
medium in which they are suspended (cf. Rhumbler, ’93). (Cf. 
also Hermann, ’89b ; Bouin, ’97 ; Mertens, ’93 ; Debski, ’97 ; 
Carnoy and Lebrun ’97a ; Koernicke, ’96.) 

7. Paranucleoli and Pseudonucleoli , Double Nucleoli , etc. 

The term paranucleolus is here adopted as equivalent to 
Flemming’s “ Nebennucleolus,” and I shall use simply the 
name “nucleolus,” or “nucleolus proper,” instead of “ Haupt- 
nucleolus.” E. B. Wilson’s terms, “principal nucleolus” and 
“ accessory nucleolus,” are somewhat inconvenient on account 
of their length, and may be misleading, since the “ principal 
nucleolus” is often smaller than the “accessory nucleolus.” 
“ Paranucleolus,” as used here, is not employed in the same 
sense as by Stuhlmann (’86), since he expresses by this term 
portions of the nuclear reticulum ; in my paper the term 
“ nucleolus ” has not been used for any part of the chromatin 
elements of the nucleus. 

In many egg cells, especially those of the Mollusca, Annelida, 
Tunicata, and Echinodermata , two kinds of nucleoli occur accord¬ 
ing to the writers on these objects, which differ from one 
another chemically and in some cases also structurally ; these 
are the nucleolus proper and the paranucleolus. Of these it 
is the nucleolus proper which seems to be morphologically 
comparable to the nucleoli of somatic cells, however the two 
may differ chemically. The paranucleolus may be either larger 
or smaller than the nucleolus, and appears usually to be distin¬ 
guishable from the latter by staining less deeply with the 
specific nucleolar stains. In the spermatoblast of the mouse 
these two kinds of nucleoli have been found by Hermann (’89) ; 
and in somatic cells by Lonnberg (’92, liver cells of Doris , Poly - 
cera, Aeolidia, and Astacus) ; perhaps the smaller of the two 
nucleoli found by me in the blood corpuscles of Doto might 
represent a paranucleolus. In plant cells apparently only one 
kind of nucleolus is present, this being comparable morphologi¬ 
cally to the nucleolus proper of the germ cells and to the nucle¬ 
oli of the somatic cells of Metazoa. Thus paranucleoli are quite 


MONTGOMERY. 


[Vol. XV. 


5*6 

frequent in many egg cells, infrequent in somatic cells of the 
Metazoa , and apparently never present in plant cells. In each 
such egg cell there may be either one nucleolus proper and 
from one to several paranucleoli (this being the most usual case), 
or there may be a single paranucleolus and a few nucleoli 
proper. In the ova of three forms examined by me there were 
two kinds of nucleoli present, namely, in Montagna, Polydora , 
and Rodalia. In my descriptions I have employed the term 
“ pseudonucleolus ” for these secondary nucleoli, since in this 
form they have a different structure from that of the nucleolus 
proper, but nevertheless stain in the same way, so it is difficult 
in this case to decide whether they correspond to paranucleoli, 
and hence I have used the indifferent name “ pseudonucleoli” 
for them. In Polydora we found from one to three paranucleoli 
in the larger germinal vesicles, and these are always apposed to 
the nucleolus. Then the smaller, deeply staining bodies in the 
maturer stages of the ovum of Rodalia may be comparable to 
paranucleoli. Whether the remarkable structures of the germi¬ 
nal vesicles of Tetrastemma catenulatum are paranucleoli, I am 
wholly unable to decide. This problem of different types of 
true nucleoli in the same nucleus is one of the most difficult 
in the study of nucleolar structures, so that it is necessary to 
discuss it more in detail. 

A. Schneider (’ 83 ), Brauer (’9i), and Floderus (’ 96 ) consider 
the paranucleoli to be derivatives of the nucleolus proper, more 
especially to be buds from its surface. Hacker (’93a) considers 
them to be secretions of the chromatin. Flemming (’ 82 ) doubts 
whether “ die Unterscheidung von Haupt- und Nebennucleolen 
eine durchgehende Geltung beanspruchen kann he finds that 
in Anodonta the two are at first in contact, but that later they 
become separated. Giard (’8i) finds in the ovum of a Spionid 
one nucleolus, and later there appears in the nucleus a much 
smaller body, which fuses with the former. Lonnberg (’ 92 ) 
thinks that the paranucleoli may serve for the acquisition of 
nourishment, or may contain reserve nourishment. List (’ 96 ) 
considers that the paranucleoli and the nucleoli of the somatic 
cells are more closely allied to one another than to the nucle¬ 
olus proper of the ova, and that the former two “ mindestens 


No. 2 .] COMPARATIVE CYTOLOGICAL STUDIES. 517 

verschiedene Modificationsstufen des Paranucleins . . . darstel- 
len.” Hessling (’54) found that in the ovum of Unio the smaller 
paranucleolus is divided off from the larger nucleolus proper. 
Hacker, in his last paper on the subject (’95), considers that the 
paranucleoli are of later formation than the nucleolus proper. 

Now in many of those cases where a paranucleolus and a 
nucleolus have been described lying in contact with one another 
it is very probable that the vacuolar portion of the vacuole has 
been described as a paranucleolus. I have no doubt that many 
of the earlier observers, who studied the nucleolus mainly in the 
living egg, have been thus misled, since only sections of nucleoli 
can show the true nature of the nucleolus. Thus Lonnberg, in 
speaking of “helle Kugeln ” in the germinal spot of Mytilus , 
says : “ Es ist schwer zu entscheiden, ob es sich hier nur um 
Vacuolen handelt”; and any one studying the unsectioned nucle¬ 
olus of Montagna would be misled into supposing that here two 
nucleoli of different consistency are apposed to one another. 
Accordingly, we must be very careful in treating as facts some 
of the observations of the earlier workers, which were made 
upon unstained and unsectioned material. 

But there are undoubtedly many cases in which two kinds of 
nucleoli do occur, 1 and this is especially so in germinal vesicles. 
The nucleolus and the paranucleolus may be in contact with 
one another, may be always separated, may at first be in con¬ 
tact and later become separated, or finally may be at first 
separated and later come into mutual contact. Are these para¬ 
nucleoli derived from the nucleolus proper, or have they a 
distinct origin ? In the ovum of Polydora the paranucleoli 
appear towards the close of the maturation period, and then are 
always in contact with the outer surface of the nucleolus proper. 

1 Cf. the reviews of the following papers: Floderus (’96), Hermann (’89a, b, 
’97), Vejdovsky (’95a), Flemming (’74, ’82), Hacker (’93a), Kultschitzky (’88), 
Lukjanow (’87b), Brauer (’91, ’92), Nussbaum (’87), Rein (’83), Henking (’87), 
Van Beneden (’80), Leydig (’55a, ’50), Stauffacher (’93), Stepanoff (’65), Giard 
(’81), Mark (’77, ’81), Lonnberg (’92), Stuhlmann (’86), List (’96), Van Bemme- 
lin (’83), Platner (’86), Clapar£de (’69), Hessling (’54), Ruckert (’94), Bouin (’97), 
Vom Rath (’95b), Moore (’95), Weismann and Ishikawa (’89), Fol (’89), Lacaze- 
Duthiers (’57), Fauvel (’97), Held (’95), Michel (’96), Steinhaus (’88), Metzner 
(’94), Braem (’97), Siebold (’39), Reinhard (’82), Kraepelin (’92), Davenport 
(91). 


MONTGOMERY. 


[VOL. XV. 


518 

In the ova of Montagna and Rodalia they are never in contact 
with the nucleolus. In none of these three cases observed by 
me does there seem to be any genetic connection between the 
paranucleoli and the nucleoli proper. And in other cases, 
where the two are separated (this separation is the most usual 
state), no genetic connection between the two has been de¬ 
scribed ; and even in that smaller number of cases where 
they are in contact with each other at some period of their 
development, no positive proof of their genetic relation has 
been offered. Therefore we might conclude, though with re¬ 
serve, that in the greater number, if not all, cases the para¬ 
nucleoli are not derivatives of the nucleolus, but are products 
sui generis. It is the rule that the nucleolus proper appears 
in the nucleus before the paranucleoli arise, the latter usually 
arising first towards the close of the growth stages. Accord¬ 
ingly, though I cannot corroborate Hacker’s (’95) conclusions 
as to the origin of the nucleolar substance, I am inclined to 
agree with him that portions of nucleolar substance are succes¬ 
sively deposited in the nucleus, and that those portions which 
are deposited last, after the nucleus has undergone important 
physiological and chemical changes, would differ from the 
portion first produced (that of the nucleolus proper), and so 
would represent the paranucleolus. And there are certain facts 
from my own observations which would support this view. In 
the earlier stages of the maturation of the ovum of Tetrastemma 
and Zygonemertes there are a large number of nucleoli produced 
successively at the periphery of the nucleus; these then wander 
successively to the center of the nucleus, and then from that 
point again to the periphery. Now in this last stage, when the 
nuclear filaments are commencing to arise, we find, usually in 
contact with the latter, much smaller, more deeply stained 
nucleoli, and these I have termed “nucleoli of the second 
generation.” We have found, accordingly, that after the nu¬ 
cleus has passed through very marked physiological changes 
(increase in size, redistribution of chromatin), another kind of 
nucleoli appears, which may or may not be morphologically com¬ 
pared to the paranucleoli of other ova. These nucleoli of the 
second generation have neither a genetic nor a physiological 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 519 

relation to those of the first generation ; and their difference 
from the latter is probably due to the fact that they have been 
produced at a time when very different physiological conditions 
exist in the nucleus. 

It is not my intention in this contribution to deal in any 
detail with those cases where double nucleoli occur in a cell, or 
those where two chemically and morphologically different kinds 
of “ nucleoli” occur in the same nucleus ; to these cases it is 
my intention to devote a special study. But preliminarily, from 
those observations which I have made on this subject, the 
following conclusions are in order. In a nucleus there some¬ 
times occurs a double nucleolus, the component parts of which 
may each represent a true nucleolus ; or such a double nucle¬ 
olus may consist of a true nucleolus apposed to a chromatin- 
nucleolus (according to my unpublished observations on the 
spermatocytes of the beetle Harpalus). Further, and this is fre¬ 
quently the case in resting spermatocytes of the first order, the 
nucleus may contain a true nucleolus separated from a chromatin- 
nucleolus ; and in Pentatoma y the account of the spermatogenesis 
of which will be shortly published by me, the unique process 
occurs of the chromatin-nucleolus being a metamorphosed 
chromosome (one of the fourteen chromosomes of the last 
spermatogonic division becoming the chromatin-nucleolus of 
the first spermatocyte) ! This peculiar structure of Pentatoma 
divides with the true chromosomes in the first reduction divi¬ 
sion. In another case where I have been able to follow all 
the developmental stages of a chromatin-nucleolus, namely, in 
cells of the hypodermis of the larva of Carpocapsa ,, I found it 
to originate from one of the granules of the nuclear reticulum, 
— a particular one of these granules (karyosomes) gradually 
increasing in size until it attains large dimensions ; during its 
growth period it is usually attached to one of the true nucleoli 
of the cell. What is of importance in these two cases (Penta¬ 
toma and Carpocapsa) is the distinction emphasized between the 
true nucleolus and a karyosome or chromatin-nucleolus : the 
latter always standing in genetic connection with the true 
chromatin, while the former, so far as my observations go, is 
never derived from this substance. These observations are not 


520 


MONTGOMERY. 


[Vol. XV. 


wholly out of place in the present paper on the true nucleolus, 
since they are necessary to prove that the true nucleolus is in 
all cases never derived from the chromatin ; where “ nucleoli ” 
have been described as arising from the chromatin elements of 
the nucleus, such structures cannot correctly be included under 
the term “ nucleolus,” when the latter is used in the proper 
sense. 


8. Relation between Nucleoli and Centrosomes. 

The greater number of cytologists agree that there is no 
genetic relations between these two structures ; and my obser¬ 
vations on the egg of Piscicola as well as more recent studies on 
other objects corroborate this view. But some few have been 
led to contrary conclusions by observing the fact that in mitosis 
the nucleolus often disappears about the time that the centro- 
some becomes apparent. Thus Karsten (’ 93 ) assumes that the 
nucleoli wander out of the nucleus into the cytoplasm, and 
there become the centrosomes of the spindle ; this observation 
has been refuted by Humphrey (’94). Also Wasielevsky (’ 93 ) 
believes that the centrosomes of the egg of Ascaris stand in 
some connection to the nucleoli, but this stands in direct oppo¬ 
sition to the conclusions of all other workers on this object, 
except those of Carnoy and Lebrun (’97b), and the supposition 
of Sala (’95). Then Lavdowsky (’94) concludes that the nucleo- 
linus is the centrosome in the process of formation, but he failed 
to observe the steps by which this body develops into a centro¬ 
some. Further, Julin (’93b) is said by Delage (’95) to have 
assumed a genetic relation between the centrosome and the 
nucleolus. Other supporters of the nucleolar origin of the cen¬ 
trosome : Balbiani (’ 95 ), Wilcox (’95), Bremer (’95b), F. Toyama 
(’ 94 ). I believe that these are the only investigators who have 
assumed this genetic relation. We may conclude, from the 
greater number of observations at hand, that there is probably 
no connection between these structures in the metazoan cell. 
But it is difficult to decide the homologies of the body found by 
Keuten (’95) in the nucleus of Ceratium , and termed by him 
nucleolo-centrosoma ; he considers it as equivalent to the central 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


521 


spindle and centrosome of Ascaris, but might it not be com¬ 
pared to the nucleolus alone, or to the nucleolus plus centro- 
somes of the metazoan cell ? However, the significance of 
most protozoan “ nucleoli ” is very problematical. ( Cf . the later 
observations of Lauterborn, ’95a.) 

9. Ontogenetic Origin of the Nucleolus. 

Very few observations have been made to determine the 
mode of origin of the nucleolus, though there are numerous 
hypotheses intended to explain it. We may leave aside, for the 
time being, its mode of reappearance in the daughter-nuclei 
after nuclear division, since a special section will be devoted to 
that subject. 

In order to determine the mode of origin of the nucleolus in 
resting stages of nuclei, I have studied those cells in which at 
first no nucleolus is present, but which after a certain period of 
growth acquire one. Objects well adapted for such investiga¬ 
tion are the ova of the nemerteans and the mesenchym cells 
of Cerebratulus. For details of these processes the reader is 
referred to the observations. 

In the ova of the nemerteans the nucleoli at the time of their 
first appearance are always in close contact with the nuclear 
membrane ; this is also the case for the mesenchym cells of 
Cerebratulus , and probably for the paranucleoli of the ova of 
Rodalia. In all these cells the nucleoli only then leave the 
periphery of the nucleus and wander towards its center, after 
the nucleus has increased more or less in size. There is only 
one explanation for the peripheral position of the nucleoli at the 
time of their first appearance, namely, that their substance is 
extranuclear in origin. This process of formation has already 
been discussed in detail for the several cells, and it is not 
necessary to repeat here all the detailed observations on which 
the main deduction is based. If the nucleolar substance were 
a secretion of the nucleus, as Hacker (’ 95 ) assumes, how would 
this assumption explain the strictly peripheral position of the 
nucleoli when they first arise? For on Hacker’s hypothesis 
we should expect the supposed nucleolar secretions to be de- 


522 


MONTGOMERY. 


[Vol. XV. 


posited evenly throughout the nucleus, and not only at the 
periphery. And his deductions are based in great part, as those 
of most other investigators, on the study of maturation mitoses, 
and he had not observed their first mode of origin, namely, their 
origin in nuclei which are not in the prophases of mitosis, but 
are only gradually becoming differentiated from somatic cells. 
I have found no evidences in any cell that the nucleoli stand 
in any genetic relation to the chromatin elements of the 
nucleus ; and while the chromatin may derive substances from 
the nucleoli, I am unacquainted with any observations which 
show that the nucleoli derive any part of their substance from 
the chromatin. In all the cases observed by me, the nu¬ 
cleus appears to assimilate a substance or substances from the 
cytoplasm, and after this substance has entered the nucleus it 
apparently undergoes there a chemical change, and becomes 
deposited on the inner surface of the nuclear membrane in the 
form of masses of varying dimensions, which may be either 
globular or irregular in shape, according as they are fluid or 
viscid in consistency. In the case of the ova of the nemerteans 
the substance taken up into the nucleus, and which there 
becomes deposited in the form of nucleoli, is sometimes exactly 
similar to the substance of the yolk-balls which lie in the cyto¬ 
plasm; in other cases it is probably similar to those metabolically 
changed portions or inclusions of the cytoplasm, out of which 
the yolk-balls are later differentiated. In Linens , indeed, the 
yolk-balls may often be found halfway through the nuclear mem¬ 
brane, and their appearance is exactly similar to that of the 
nucleoli. In the mesenchym cells of Cerebratulus the substance 
of the nucleoli appears to be identical with that of the numer¬ 
ous nutritive granules which are dispersed in the cytoplasm ; 
the latter globules arise in the cytoplasm before the nucleolus 
appears in the nucleus, and as soon as they become numerous 
in the neighborhood of the nucleus, peripheral nucleoli begin to 
appear in the latter. In the subcutical gland cells of Piscicola 
the nucleolus, at the time of its most rapid growth, is apposed 
to the nuclear membrane; but when this period of volume- 
increase has ceased, it is never found in this position. Further, 
the paranucleoli of Rodalia appear first in contact with the 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 523 

nuclear membrane. Schwalbe (’ 76 ) found in the nuclei of 
various vertebrate embryos that when the nucleoli first arise 
they appear as thickenings of the inner surface of the nuclear 
membrane. 

From these observations I conclude, accordingly, that the 
nucleolar substance, in many if not all cells, has an extranu- 
clear origin ; and that, though it may undergo a chemical change 
after entering the nucleus, it can be regarded neither as a secre¬ 
tion nor as an excretion of the latter. In making this con¬ 
clusion I can corroborate the views of only one investigator, 
namely, Korschelt (’ 89 ), though he changed this opinion in a 
later paper (’ 97 ). He concluded that the nucleolar substance 
stands in some connection with the nutritive processes of 
the cell, and that the nucleus probably derives it from the 
cytoplasm. 

Other views on the origin of the nucleolus (those of Hacker 
have already been mentioned): Auerbach (’ 74 a, ’ 76 ) first supposed 
the nucleolus to be cytoplasmic in origin ; more recently (’ 90 ) 
he appears to champion its nuclear origin. Rhumbler (’ 93 ) 
assumes that the “ Binnenkorper ” of Protozoa are products of 
the nucleus, but he does not attempt to decide whether those 
of the Metazoa have a similar origin. Strasburger (’ 82 b) also 
postulates a nuclear origin for the nucleolus, and assumes that 
its substance is allied to chromatin. Jordan (’ 93 ) holds that 
the nucleoli probably arise from the chromatin threads. Flem¬ 
ming (’ 82 ) considers them to be “ specifische Produkte des 
Kernstoffwechsels.” Schwalbe (’ 76 ) supposes the nucleolar 
substance to be at first identical with that of the nuclear mem¬ 
brane, since he found it to arise as thickenings of the latter. 
C. Schneider (’ 91 ) supposes it to be a metamorphosed portion 
of the chromatin. Leydig (’ 83 ) concludes that the nucleoli are 
portions of the chromatin reticulum. Guignard (’ 85 ) assumes 
that they are derivatives of the chromatin filaments. Watase 
(’94) considers them to be metabolic products of the cell, but 
he gives no detailed observations in regard to their mode of 
formation. Mertens (’93) and Retzius (’ 81 ) consider them to 
arise by concentration of the chromatin reticulum. 


524 


MONTGOMERY. 


[Vol. XV. 


io. Discharge of Nucleolar Substance from Resting Nuclei. 

Will (’84) holds that the larger nucleoli of the amphibian ger¬ 
minal vesicle pass out into the cytoplasm, and there become the 
yolk-nuclei ; and Scharff (’88) corroborates this view for the ova 
of Trigla , though it is opposed by Cunningham (’95). Macallum 
(’9i) concludes that in amphibian ova the peripheral nucleoli 
generate a substance which diffuses first in the nucleus and from 
there into the cytoplasm, and that this substance combines with 
the cytoplasm to form the yolk substance ; Jordan (’93) expresses 
a somewhat similar view in regard to the yolk formation of the 
newt. Henneguy (’93) assumes that the corpuscle of Balbiani 
in the ova of Vertebrata “ est tres probablement une partie de 
la tache germinative, ou une tache germinative entiere, qui sort 
de la vesicule [germinative] pour penetrer dans le vitellus,” and 
Mertens (’93) holds a similar view. And for egg cells of 
Tunicata , Floderus (’96) confirms Roule’s (’84) observations, 
that the “ intravitelline Korper ” are paranucleoli which have 
wandered into the cell body. Cf. also Bremer (’95a, b). 

Leydig (’88) finds that in ova of Geophilus , Stenobothrus, 
Rana, and Triton particles of nucleolar substance penetrate 
into the cytoplasm. Lukjanow (’88) concludes that in the case 
of the cells of the stomach mucosa of Salamandra , the nucleo¬ 
lus discharges a portion of its substance from the nucleus. 
Humphrey (’94), from observations on plant cells, maintains 
that in some cases portions of nucleolar substance may pass 
into the cytoplasm. 

Fol (’83a, b) concludes that the follicle cells of the ascidian 
egg arise as buds from the surface of the germinal vesicle, 
and that each of these buds contains a particle of nucleolar 
substance ; these conclusions are affirmed by t Roule (’83). 
Scharff (’88) supposes that the follicle cells of the ovum of 
Gadus are derived from nucleoli which have left the germinal 
vesicle, such nucleoli becoming the nuclei of the new cells. 
(Ogata ’83) studied human pancreas cells and finds that a 
nucleolus wanders out of the nucleus, becomes a “ Nebenkern,” 
and the latter finally changes into the nucleus of a new cell, a 
conclusion which is opposed by Platner (’89b). 


No. 2 .] COMPARATIVE CYTOLOGICAL STUDIES. 


I have found a wandering of nucleolar substance out of rest¬ 
ing nuclei in one very beautiful and unique case, namely, in the 
subcuticular gland cells of Piscicola; at one stage in its cycle 
of development the nucleus commences to contract in volume, 
and in so doing discharges all except a single one of its nucleoli 
into the cytoplasm. This and certain of the observations cited 
from other investigators show that a discharge of nucleolar 
substance from the resting nucleus takes place in some cells. 
But the more recent observations of Morgan, Floderus, and 
others on Tunicate development render it very probable that 
Fol and Roule were mistaken in assuming that the nucleoli 
which pass out of the germinal vesicle become the constituents 
of follicle cells. There is still some question, also, as to whether 
the nucleolar substance in the cytoplasm takes any part in the 
formation of the yolk substance. Other pertinent observations : 
Mertens (’93), Bremer (’95a, b), Kosinski (’87, ’93), Galeotti (’95), 
Melissinos and Nicolaides (’90), Auerbach (’74), Ver Ecke 
(’93), Steinhaus (’88), Rohde (’96). 

11 . Behavior of Nucleoli during Nuclear Division. 

It is in cases of nuclear division that the nucleolus has 
received the most attention from morphologists. The behav¬ 
ior of the nucleolus in mitosis and amitosis may be treated 
separately. 

i. Amitosis. — In this mode of nuclear division it is frequently 
the case for the nucleolus to divide first, so that each of the 
daughter-nuclei receives a half, or approximately a half (for the 
division of the nucleolus is not always into two equal parts), 
of the parent-nucleolus. In support of this deduction the fol¬ 
lowing observations may be mentioned : Schaudinn (’94, Amoeba 
crystalligera)\ F. E. Shulze (’75, A. polypodia)\ Will (’85, ova of 
Nepa , Notonecta) ; Doflein (’96, degenerating ova of Tubularia) ; 
Carnoy ('85, ova of Gryllotalpa y Lithobius , Geotrupes ); Korschelt 
(’95, intestinal cells of Ophryotrocha ); my observations on the 
peritoneal cells of Polydora; Hoyer (’90, intestinal epithe¬ 
lium of Rhabdoncma ); Frenzel (’93b, hepatopancreas cells of 
Astacus ); Platner (’89a, Malpighian tubes of Dytiscus ; Wheeler 


526 


MONTGOMERY. 


[Vol. XV. 


(’89, follicle cells of Blatta)\ de Bruyne (’97, follicle cells of 
Nepa , Periplaneta , Meconema, Aeschnd). E. B. Wilson (’96), 
in speaking of amitosis, states : “ In many cases, however, 
no preliminary fission of the nucleolus occurs ; and Remak’s 
scheme must therefore be regarded as one of the rarest forms 
of cell division.” But the list of cases which I have given 
shows that such cases of nucleolar division are frequent in 
amitosis, so that I conclude that a fission of the nucleolus, if 
not exactly typical for this mode of nuclear division, is never¬ 
theless well represented and occurs here much more frequently 
than in mitosis. Dr. E. G. Conklin has demonstrated to me 
preparations of nucleolar division in follicle cells of Gryllus , 
which he has kindly allowed me to mention here. 

2 . Mitosis. — In karyokinesis the nucleolus may either not 
disappear, or, and this is the most usual case, it disappears 
before the spindle is formed. These two modes may be con¬ 
sidered in turn. 

(a) The nucleolus does not disappear. — In some few cases 
the nucleolus wanders out into the cytoplasm after the disap¬ 
pearance of the nuclear membrane and may remain there for 
some time without undergoing any change. Such cases have 
been described by Hacker (’92a, egg of Aequorea ), Wheeler (’95, 
that of Myzostoma ), H. V. Wilson (’94, ova of Tedamione and 
Hircinia), Tangl (’82, flower buds of Hemerocallis ), Gjurasin 
(’93, Peziza ), and Karsten (’93, sporangia of Psilotum). In all 
these cases the nucleolus ultimately disappears in the cytoplasm, 
though in Aequorea it may be observed still in the cell body of 
one of the blastomeres at the thirty-two cell stage, and the 
daughter-nuclei produce their own nucleoli. (Similar are the 
observations of Mead, ’95; Hacker, ’96, ’97; Rosen, ’95; Zimmer- 
mann, ’96; Metzner, ’94; Foot ’94; Poirault and Raciborski,’96.) 

In the other cases where the nucleolus does not disappear it 
remains within the nucleus. In some of these cases it appears 
to divide into two or more parts; in other cases it may be that 
one of the daughter-nuclei receives the whole parent-nucleolus, 
while in the other one a new nucleolus is produced. There are 
a few observations which show that it sometimes divides ; thus 
Strasburger (’82b, embryo sac of Galanthus) and Rosen (’92b, 


No. 2 .] COMPARATIVE CYTOLOGICAL STUDIES. 527 

Sync/iytiiuni) ; Reinke studied the mitosis of the spleen cells 
of the mouse, and found that the single parent-nucleolus divides 
into three or four pieces, while at the end of the mitosis each 
daughter-nucleus contains a single nucleolus. In the mitoses 
of the ovogonia of Linens and Polydora my own observations 
show that the nucleolus persists in the nucleus, and each 
daughter-nucleus contains one nucleolus, so that it is very 
probable that in these cases the parent-nucleolus divides into 
two, and each daughter-nucleus thereby receives a half of it ; 
but these mitoses were so small that I was unable to decide 
this point definitely. Rosen (’95) finds nucleolar division in 
root cells of Phaseolus ; J. Wagner (’96a) describes a similar 
division of a “ nucleolus ” in spermatocytes of Arachnids, though 
this case, like that described by Henking (’90), probably repre¬ 
sents a chromatin nucleolus. This persistence of the nucleolus 
in the nucleus during mitosis must be considered atypical. 

(b) The nucleolus disappears during mitosis. — This is the 
most usual mode of behavior of the nucleolus during mitosis. 
The nucleolus either gradually diminishes in size, and so finally 
vanishes, or else it first fragments into a number of smaller 
pieces, and then these disappear. The only cell which I had 
for the study of this phenomenon was the ovum of Piscicola 
during the formation of the first pole spindle. When this 
spindle is complete no trace of nucleolar substance is to be 
seen anywhere in the cell. In stages immediately antecedent 
to that of the spindle, numerous minute granules, as well as a 
smaller number of larger globules, are dispersed through the 
nuclear sap; all these stain with eosin, and I regard them as 
particles of nucleolar substance which had become separated 
from the nucleolus. Thus a dissolution of the nucleolar sub¬ 
stance commences before the nuclear membrane has disap¬ 
peared, and after this membrane has vanished it is probable 
that all the nucleolar substance must be dissolved by the action 
of the cytoplasm, or at least become dispersed through the 
latter, so that no remnant of it is to be found in the region of 
the spindle or of the chromosomes. During the process of 
dissolution of the nucleolar substance in the nuclear sap the 
chromatin elements stain red (with eosin), and this fact may be 


MONTGOMERY. 


[Vol. XV. 


528 

explained either by the assumption that the nucleolar substance 
unites chemically with the chromatin, or that it simply pene¬ 
trates into the meshes of the latter ; since no nucleolar 
substance appears to be united with any of the twelve chromo¬ 
somes we may conclude that it does not unite chemically with 
the chromatin, and therefore the chromosomes probably do not 
serve to carry it over into the daughter-nuclei. We may now 
briefly review the results of other observers on the mode of 
disappearance of the nucleolus during mitosis. 

It is not necessary to discuss the earlier view of O. Hertwig, 
which he has since discarded, that “ der Eikern der aus dem 
Keimblaschen frei gewordene oder ausgewanderte Keimfleck 
ist,” nor yet the view of Kolliker. Kleinenberg (’ 72 ) believes 
that the germinal spot of Hydra dissolves during mitosis ; 
Brauer (’91) finds that it breaks into fragments, of which a part 
seems to be dissolved in the cytoplasm, “ ein Theil tritt unver- 
andert nach dem Schwinden der Membran in das Eiprotoplasma 
liber.” Fick (’93, germinal spot of Amblystoma) finds that the 
nucleoli disappear at the time of the longitudinal splitting of the 
chromosomes ; and Bohm (’88) reaches the same conclusion for 
Petromyzon. Davidoff (’89, ovum of Distaplia ) concludes “ dass 
aus dem Nucleolus ein Kern mit Kernnetz, mit einem Nucleolus 
und Nucleolinus hervorgegangen ist”; and Vejdovsk^ (’88, 
Rhynchelmis ), Blochmann (’82, Neritina ), and Marshall (’92, Gre- 
garina ) conclude that the nucleoli become chromosomes. In the 
egg of Ascaris the nucleoli gradually disappear, according to most 
observers. Strasburger (’82b) first contended that the nucleolar 
substance is taken up into the nuclear filaments ; later (’88) he 
writes : “ Auf Grund meiner neueren Erfahrungen erscheint es 
mir iiberhaupt unwahrscheinlich, dass die Nucleolarsubstanz, 
auch nach ihrer Auflosung im Kernsafte, den Kernfaden als 
Nahrung dienen sollte,” and he considers that after it is 
dissolved in the nuclear sap a portion of it forms the cell 
membranes of the daughter-cells (cf. also his paper of ’93). 
Rein (’83, ova of Lepus and Cavia) finds that the nucleolus 
breaks into small fragments, which finally disappear in the 
substance of the nucleus. Pfitzner (’83, ectoderm cells of 
Hydra) terms the nucleolar substance “ prochromatin,” since 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


529 


he finds that in mitosis it changes into chromatin. Rabl (’85, 
larval cells of amphibians) and O. Schultze (’87, ova of Rana 
and Triton) contend that the nucleolar substance takes some 
part in the formation of the nuclear filaments ; but Born (’ 94 ) 
subsequently found that these filaments stand in no connection 
with the nucleolar substance. Holl (’93, ovum of Mus ) finds 
that the central granules of the nucleoli wander out of them 
and so become the chromosomes. Van Beneden (’ 75 , ovum of 
Lepus) originally supposed that the nucleolus becomes the first 
pole body. Kastschenko (’90, ova of Selachii ) finds that all the 
nucleoli disappear in the spirem stage, while Rtickert (’92) 
finds that a few of them pass into the cytoplasm. Stuhlmann 
(’86, ova of Insecta ) finds that the nucleoli gradually disappear 
during the maturation of the egg ; and similar conclusions 
were reached by Stauffacher (’93, Cyclas), Rhumbler (’95, 
Cyphoderia ), Sheldon (’90, Peripatus)> Heathcote (’86, Julus ), 
Van der Stricht (’95, Amphioxus ), Brauer (’92, Branchipus ), and 
Vejdovsky (’82, Sternaspis). Auerbach (’96, spermatogonium 
of Paludina) holds that the nucleolar substance becomes incor¬ 
porated with the chromatin elements. Meunier (’86) and Moll 
(’93) for Spirogyra , and Carnoy (’85) for other cells also, hold 
that the chromosomes are derivatives of the chromatin skein of 
the nucleolus. Heuser (’84, mitoses of various plant cells) con¬ 
tends that the nucleoli become gradually apposed to the nuclear 
filaments, and that their substance unites with these elements, 
though in some cases a superfluous portion of the nucleolar sub¬ 
stance may be discharged from the nucleus. Korschelt (’95, 
ovum of Ophryotrocha) finds that the nucleolus gradually dis¬ 
appears by dissolving in the nuclear sap, and believes that a part 
of this substance may be introduced into the nuclear filaments. 
Zacharias (’85) somewhat prematurely concludes that the nucleoli 
always disappear in mitosis. Tangl (’82) finds that in Hemerocal- 
lis , in uninucleolar nuclei, the nucleolus dissolves in the nucleus, 
but in those which are multinucleolar one may pass out into the 
cytoplasm ; in Hesperus and Cisium they gradually disappear. 
Humphrey (’94, plant cells) holds that “die Nucleolen in einigen 
Fallen aus der Kernhohle, bevor sie von den karyokinetischen 
Kraften angegriffen werden, austreten konnen.” 


530 


MONTGOMERY. 


[Vol. XV. 


Wager (’93, Agaricus ) describes the nucleoli as becoming 
dissolved in the caryolymph, and then, this dissolved substance 
penetrating the chromatin elements, the latter serve to carry 
it over into the daughter-nuclei. Went (’87, plant cells) holds 
“ dass in vielen Fallen wenigstens der Nucleolus beim Anfang 
der Kerntheilung im Kernfaden aufgenommen wird,” and that 
“er sich nach der Theilung auch wieder daraus bildet.” 
Riickert (’94, egg of Cyclops ) finds that the nucleoli gradually 
break into fragments and the latter disappear. But there is 
not space here to mention all the views of students of mitosis. 

There are only a few observations which would show that in 
mitosis the chromosomes are derived from the nucleoli (David- 
off, Vejdovsky, Blochmann, Marshall, Sobotta, ’95, Macallum, 
'95, Carnoy, ’97a, R. Hertwig, ’96, not corroborated by Brauer, 
’94), and these cases stand in such marked contradiction to the 
observations of other morphologists that a reinvestigation of 
them is very necessary. 1 Then we have the observations of 
Carnoy, Meunier, and Moll, which would show that the chro¬ 
mosomes are derived from a part of the nucleolus ; but the 
existence of a “ nucldole-noyau,” i.e ., of a nucleus within a 
nucleus, as assumed by Carnoy and his followers, in any meta¬ 
zoan cell, seems to be very problematical. On the other hand, 
most observers agree that the nucleoli disappear more or less 
gradually during mitosis, and that the chromosomes are not 
derived from them. Now we have reached the crucial ques¬ 
tion : What is the mode of transference of the nucleolar 
substance to the daughter-nuclei ? In answer to this, some 
observers hold that this substance may be distributed in the 
cytoplasm and taken up therefrom into the daughter-nuclei; 
others, that it combines with the chromatin elements and is 
transferred with these; still others maintain a position inter¬ 
mediate between these two. 2 But when we find so much vari¬ 
ance in the conclusions of competent investigators only one 
deduction is allowable, namely, that the mode of transportation 

1 On the relation of nucleoli to chromosomes, cf. also Cunningham (’97), 
Sobotta (’95), Macallum (’95), Platner (’89c), Carnoy (’97a), R. Hertwig (’96), 
Van Beneden (’83), Zimmermann (’96), Lauterborn (’96), Boveri (’88), Wheeler 
(’97). 

2 Cp. also Belajeff (’94), Mottier (’97), and Rosen (’95). 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 531 

of the nucleolar substance is probably different in different 
objects. 

We have found above that in the simplest though secondary 
nuclear divisions, the amitotic, the nucleolar substance of the 
parent-cell is transported into the daughter-nuclei by the me¬ 
chanically simplest process, namely, by a direct division of the 
parent-nucleolus ; this is very frequently the case in amitosis, 
though it does not always occur. But in most mitotic divisions 
the nucleolus first disappears, i.e. y there would seem 'to be an 
indirect mode of transference of its substance corresponding to 
the indirect mode of transference of the chromatin and linin ele¬ 
ments. Now all mitotic divisions do not proceed on exactly the 
same plan, for we find differences in regard to the presence of a 
central spindle, in regard to the number of the chromosomes, etc. 
Accordingly, one would expect also different modes of transfer¬ 
ence of the nucleolar substances. Thus in some cases, as 
Wager (’93) suggests, the chromosomes may serve as mechan¬ 
ical vehicles for the transportation of this substance. In many 
other cases it is very probable that this substance, after the 
disappearance of the nuclear membrane, becomes dispersed in 
the cytoplasm ; and then each of the daughter-nuclei may 
either take up this substance from the cytoplasm again, or may 
produce its own nucleolus from a new substance, owing to the 
primitive nucleolar substance having been assimilated by, or 
even discharged from, the cytoplasm. There are observations 
in support of each of these three modes of re-formation of 
nucleoli in the daughter-nuclei. But since when the nuclear 
membrane disappears the cytoplasm probably comes into con¬ 
tact with the substance of the nucleoli, it is most probable that 
it would produce either a physical or a chemical change in the 
latter, and hence the second and third modes would appear 
the more probable. Accordingly, I agree with Humphrey (’94) 
that there is no substantial basis for Zimmermann’s (’93) con¬ 
clusion “omnis nucleolus e nucleolo,” or more strictly speaking, 
that the nucleolus in most cases is not derived from a previously 
existing one. But the third mode of diffusion of the nucleolar 
substance is in reality not a transference of this substance at 
all, since it probably becomes lost in the cytoplasm ; and hence, 


532 


MONTGOMERY, 


[Vol. XV. 


though the mode of disappearance of this substance may be 
more or less dependent upon the mode of mitosis, the substance 
of the parent-nucleolus may be in many cases not transferred 
to the daughter-nuclei, but the latter (perhaps as a rule) may 
produce their own nucleoli de novo. 

Strasburger (’93, ’97) assumes that the small granules found 
by Kostanecki (’92) in the equatorial plate of the central spindle 
may be nucleolar particles, and accordingly that the nucleolar 
substance may be in this way very evenly distributed to the 
daughter-nuclei; but it is not as yet clearly shown that these 
granules are derivatives of the nucleolus (cf also Debski, ’97, 
Sala, ’95, Pfitzner, ’86b, and Rosen, ’95). 

Zacharias (’85), Carnoy (’85), and Platner (’86) have concluded 
that in some cases the achromatic spindle fibers are derived 
from the nucleolus ; similar views are held by Strasburger (’95, 
’97), Harper (’97), and Fairchild (’97), but most facts would show 
this view untenable. 

Rhumbler (’93) assumes that a greater amount of nucleolar 
substance is accumulated in the nucleus before mitosis than is 
necessary for its growth, and this superfluous amount would 
serve for the formation of the nucleoli in the daughter-nuclei. 


12 . The Function of the Nucleolus. 

The attempt to deduce the physiological economy of a struc¬ 
ture from a mere study of its morphological relations is always 
difficult, and this is especially the case with regard to the 
nucleoli. 

Balbiani (’64) found contractile and discharging vacuoles in 
the germinal spot of Phalangium, and notes that they differ 
from the contractile vacuoles of the Rhizopoda in that they 
are not formed again at the same point. Hacker (’93c) regarded 
the nucleolus of the ovum of Echinus as an excretory organ, 
since he found its large vacuole to be contractile; he compared 
it directly to the contractile vacuole of Infusoria. Balbiani 
(’65b) also observed contractile vacuoles in the germinal spots 
of Helix , Vortex , and Prostomum , and in these the vacuole dis¬ 
charges through a small orifice in the cortical substance of the 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


533 


nucleolus. Bohm described (’88) the vacuole of the germinal 
spot of Petromyzon as connected by a fine duct with the sur¬ 
face of the nucleolus. Lukjanow (’88) found in the stomach 
cells of the salamander that the nucleolus is apposed to the 
nuclear membrane, through which it discharges an excretion. 
Compare also Van Bambeke (’97a) and Michel (’96). These 
observations would show that the nucleolus in some cases con¬ 
tains a contractile vacuole, and that the fluid substance of the 
latter is periodically discharged from it (cf. Hodge ’94, Van 
Bambeke, ’97, and Michel, ’96). • 

Flemming (’82) considers the nucleoli to be nuclear organs, 
and regards them either as containers or reserve supplies of 
chromatin, or as “ eine chemische Modification, Vorstufe oder 
Doppelverbindung ” of the latter substance; this view is also 
held by Van Bambeke (’85). Zacharias (’85) also thinks that 
they are organs, but does not agree with Flemming that they 
are reserve masses of chromatin; Gjurasin (’93) corroborates 
the views of Zacharias. Strasburger originally contended (’84) 
that they represent reserve material, a view shared by many 
later observers ; more recently (’88) he shows that the nucleolar 
substance may play some part in the formation of the cell 
membrane, but considers that they may also have some other, 
as yet unknown, function. Korschelt (’89) concludes that they 
are formed as depositions of nutritive substances, and that their 
substance “in und vielleicht ausserhalb des Kernes zur Ver- 
wendung gebracht werden sollte.” Rhumbler (’93) assumes 
that the nucleoli (“ Binnenkorper ”) of the Protozoa represent 
“ Reservestoffe ” deposited in the nucleus and consumed in 
the growth of the latter, standing in some connection with the 
chromatin ; they are not organs, but secretions of the nucleus. 
Hacker (’95) concludes that they are not nuclear organs, but 
secretions of the nucleus formed in or from the chromatin 
elements and destined to be discharged from the nucleus dur¬ 
ing mitosis; he observes that the nucleolar substance “ein 
Stoffwechselsprodukt darstellt, dessen Erzeugung in einem 
gewissen Abhangigkeitsverhaltniss zur Intensitat der vegeta- 
tiven Leistungen von Kern und Zelie steht,” and that its amount 
stands in a direct ratio “ zur Intensitat der Wechselbeziehungen 


534 


MONTGOMER V. 


[Vol. XV. 


zwischen Kern und Zelle ”; he opposes the view “dass die 
Kernkorper aus dem Zellplasma in den Kern hineingelangen 
und hier in die Bildung des Chromatins eingehen.” Leydig 
(’85) holds that certain of the nucleoli are differentiations of 
the chromatin reticulum, others of the “ Kernplasma.” Watase 
(’94) considers that they may be metabolic products of the cell. 
Auerbach (’90) holds them to be the fundamental constituents 
of the nucleus, which is a retrogression to the earlier views of 
O.and R. Hertwig. Born (’94) states : “Die Nucleolen stehen 
in Beziehung zum individuellen Zellleben, nicht zur Fortpflan- 
zung.” Lavdowsky (’94) considers them to be reserve masses 
of chromatin. Macfarlane (’81, ’85) regards them as the tropic 
centers of the cell, and as the most important mechanical agents 
in cell division. Julin (’96b) believes they conduct the vegetal 
processes of the cell. Mottier (’97) considers the nucleolus 
“ein Kraftvorrath, welcher der Zelle nach Bedarf zur Verfii- 
gung steht ” ; and Swingle (’97), as a reserve fund of nourish¬ 
ment for the kinoplasm in mitosis. Metzner (’94) considers 
them to be of importance in the processes of mitosis (compare 
his observations). Henneguy (’93) regards the nucleolus and 
Balbianian corpuscles as corresponding with the macronucleus 
of the Infusoria (cf Julin, ’93b). These, then, are the most 
important views on the nature of the nucleolus. 1 

From my own observations the nucleolar substance would 
seem to be extranuclear in origin, and not a secretion or excre¬ 
tion of the nucleus. To be sure it may, and probably does, 
undergo chemical changes within the nucleus, but it is derived 
in the first place from the cytoplasm. I regard the nucleoli as 

1 The following list includes, I believe, all who have written on the function of 
the nucleolus : Korschelt (’89), Hacker (’93a, ’95, ’97b), O. Hertwig (’77a, ’92), 
Rhumbler (’93), R. Hertwig (’76, ’96), Fick (’93), Lukjanow (’88), Brauer (’91), 
Nussbaum (’82), Strasburger (’82b, ’84, ’88, ’95, ’97), Jordan (’93), Flemming 
(’80, ’82), Van Beneden (’75), Wasielevsky (’93), A. Schneider (’83), Henneguy 
(’93), Ruckert (’92, ’94), C. Schneider (’91), Born (’94), R. Wagner (’36, ’37), 
Auerbach (’74a), Kolliker (’43), Lonnberg (’92), Klein (’78), Macallum (’91, 
’95), Stuhlmann (’86), O. Brandt (’78), Schwarz (’87), Giugnard (’85), Macfar¬ 
lane (’81, ’85, ’92), Zacharias (’85), Watase (’94), Humphrey (’94), Gjurasin 
(’93), Mann (’92), Julin (’93b), E. B. Wilson (’96), Van Bambeke (’85), Mottier 
(’97), Swingle (’97), Rosen (’95), Metzner (’94), Wheeler (’97), Carnoy (’84, 
’86, ’97a, ’97b). 


No. 2.] COMPARATIVE CYTOLOG/CAL STUDIES. 


535 


consisting of a substance, or different substances, taken into 
the nucleus from the cell body. It seems probable, further, 
that these substances stand in some relation to the nutritive 
processes of the nucleus, and in a relation to the growth of the 
latter. Thus those nuclei which are characterized by an espe¬ 
cially large amount of nucleolar substance are growing nuclei, 
i.e.y those of egg cells in the maturation period, those of the 
subcuticular gland cells of Piscicola , the mesenchym cells of 
Cerebratulus . In the gland cells of Piscicola the volume of the 
nucleolar substance rapidly increases in amount during the 
phase of growth of the nucleus, but diminishes when the latter 
decreases in volume. Somatic cells, on the contrary, at least 
those which are undergoing no dimensional changes, contain 
a relatively small amount of this substance. It* is doubtful 
whether Hacker (’ 95 ) is quite correct in assuming that the 
amount of the nucleolar substance stands in a direct proportion 
to the intensity of the functional changes which take place 
between the nucleus and the cytoplasm; at least there are but 
few criteria to enable one to compute the degree of such an 
intensity. Thus one would suppose that in nerve cells there 
was a close and intimate correlation between nucleus and cell 
body, but the nucleoli of the ganglion cells of the nemerteans 
and Piscicola are very small. Hacker’s deduction might be 
modified as follows : where there is a close physiological rap¬ 
port ', in regard to processes of nutrition, between the nucleus 
and the cell body a relatively large amount of nucleolar sub¬ 
stance occurs in the former. 

Accordingly, we find a relatively large amount of nucleolar 
substance in growing nuclei, and hence conclude that this sub¬ 
stance stands in some connection with the processes of nutrition, 
is itself either nutritive in function or represents that portion of 
substances assimilated by the nucleus from which all nourish¬ 
ment has been extracted, and in this case it would be a waste 
product. A third possibility is that the nucleoli may represent 
accumulations of nutritive substance retained in the nucleus as 
a reserve supply; but this does not seem to be very probable, 
for by this assumption it would be difficult to explain the uni¬ 
formity in the size of the nucleoli in a given species of cell. 


MONTGOMERY. 


[Vol. XV. 


536 

It would be premature to attempt to decide the exact manner 
in which the nucleolar substance is concerned in the metabolism 
of the cell. But the facts at least show that it has an extranu- 
clear origin, and is especially abundant in growing nuclei, which 
shows that it stands in intimate connection with the phenomena 
of nutrition of the nucleus. 

Vacuoles are characteristic for certain stages in the develop¬ 
ment of many nucleoli, especially those of germinal vesicles. 
For the nucleoli of the ova of Montagna and Doto , I showed 
that the vacuolar substance is at first present in the form of 
small globules in the nuclear sap, that these become applied 
against the surface of the nucleolus, and, finally penetrating into 
the latter, represent within it the vacuoles. I was unable to 
decide the mode of derivation of the vacuoles for the other 
nucleoli studied. So in some cases this vacuolar substance 
would appear not to be a derivative of the ground substance of 
the nucleolus, but to be derived from without the latter. Thus 
such nucleoli may be considered as diosmosing structures. The 
manner of growth of nucleoli is apparently by a process of 
apposition of smaller particles of nucleolar substance to their 
surfaces, and the addition of vacuolar substance to them differs 
from this only in that the vacuolar substance is intussuscepted. 
This vacuolar substance may be also a product of the nutritive 
processes of the nucleus. 

It is a difficult question to determine whether the nucleolus 
at some stage of its development should not be considered a 
nuclear organ. In most nuclei it has a regular shape, in 
others it may be oval; in many cases the nucleolus has no 
regular shape, and in the salivary gland cells of Chironomus 
(according to Balbiani) it is convoluted. From the facts at 
hand we may conclude that the shape of the nucleolus is 
pretty constant for the particular species of cell. Now, tak¬ 
ing constancy in form as a criterion of an organ, one might 
conclude that the nucleoli are organs. But, on the other hand, 
the most frequent form of the nucleolus, namely, the spherical, 
might simply be due to its thin fluid consistency, and when it 
is more viscid in consistency its shape would be more irregular. 
Thus Rhumbler (’93) concludes that the irregular nucleoli of 


No. 2 .] COMPARATIVE CYTOLOGICAL STUDIES. 


537 


Foraminifera “ durch Zusammenfliessen anfanglich leicht fliis- 
siger, dann zahfliissiger und schliesslich erstarrender Massen 
entstanden sind.” It may be asked : Why does the nucleolus 
persist through the whole resting state of the nucleus if it be 
not an organ ? It may be simply stored in the nucleus until at 
the time of mitosis, when the nuclear membrane disappears, 
it has an opportunity to leave the nucleus. The only observa¬ 
tions which would prove that the nucleolar substance may 
functionate as an independent organ are those according to 
which the nucleolus contains a contractile vacuole, and thus 
rhythmically contract and expand ; in these cases the nucleolus 
might be regarded as a pulsating excretory organ of the nucleus. 
The hypothesis might be suggested that though the nucleolus 
probably consists of substances which stand in some relation to 
the nutritive processes of the nucleus, and so at the time of its 
first formation may be a functionless, inert mass of substance, 
yet it may at later periods in the history of the resting nucleus 
acquire some active function and thus gradually come to 
acquire the value of a nuclear organ ; this hypothesis is put 
forward merely as a tentative one. According to this view the 
nucleolus might be considered as an organ which serves to 
accumulate in itself the waste products of the nucleus, thus 
serving as a reservoir for such substances ; or it might be con¬ 
sidered as an organ of excretion, to discharge waste products 
out of the nucleus : in either case the nucleolus would seem to 
stand in direct connection with the nutritive substances and 
forces of the nucleus. 


13. Comparison of the Nucleoli in Plants , Protozoa , and 
Metazoa. 

I have made no morphological studies on the nucleoli of 
plant cells, but would judge from the results of botanical inves* 
tigators that they are probably strictly comparable to the 
nucleoli of the metazoan cells. 

Rhumbler (’93) doubts whether the nucleoli of the Metazoa 
and the “ Binnenkorper ” of the Protozoa are homologous 
structures ; and, indeed, there are certain nucleolar structures 


538 


MONTGOMERY. 


[Vol. XV. 


in Protozoa which are unique, such as the nucleolo-centrosome 
of Keuten (’ 95 ). Henneguy considers that the corpuscle of 
Balbiani, together with the nucleolar elements of the metazoan 
cell, corresponds to the macronucleus of the Inftisoria ; in con¬ 
nection with this view may be mentioned the observations of 
Butschli (’80), according to which only the macronuclei of the 
Ciliata contain nucleoli. Henneguy’s hypothesis is very ingen¬ 
ious, and opens an interesting field for investigation, but it is 
difficult to determine whether it corresponds to the facts at 
hand, or whether it does not . 1 Some of the nucleoli of Protozoa 
are comparable to those of Metazoa , but it is doubtful whether 
all of them are . 2 Thus it may be the case in some of the 
gregarines that the chromatin (or its physiological equivalent) 
is localized in some or all of the nucleoli, and such structures 
could not be compared with the nucleoli of the metazoan cell. 

As to the metazoan nucleoli, there is the question whether 
the nucleoli of egg cells and of somatic cells should be consid¬ 
ered homologous. In my opinion this may be answered in the 
affirmative, since the nucleoli of both kinds of cells appear to 
be depositions of substances which are concerned in the nutri¬ 
tive processes of the nucleus. In making this conclusion I 
limit myself to the true nucleoli and do not consider those 
structures which have been erroneously termed nucleoli, but 
which in reality are portions of the chromatin reticulum of the 
nucleus. Numerous writers have considered the thickened 
nodal points of the nuclear network to be nucleoli, and here 
may be mentioned Leydig, Klein, Waldeyer, and others. The 
“ cyanophilic ” nucleoli of Auerbach (’90), the “ pseudonucleoli ” 
of Rosen (’92a), the “ nucleoles nucleiniens ” of Carnoy (’85), 
and the “ Karyosomata ” of Ogata (’83), Lukjanow (’87b), and 
Macallum (’ 91 ) are undoubtedly not nucleoli but portions of 
the nuclear reticulum. While the “ erythrophilic ” nucleoli 
of Auerbach, the “ Eunucleoli ” of Rosen, the “ nucleoles 

1 On the genetic relation of nucleoli to Balbianian corpuscles (true yolk-nuclei), 
a relation which seems to me very doubtful, cf. Mertens (’93), Galeotti (’95), 
Melissinos and Nicolaides (’90), Weismann and Ishikawa (’89), Ver Ecke (’93), 
Steinhaus (’88), Henneguy (’93), Julin (’93b). 

2 For the central masses of chromatin found in many protozoan nuclei, Doflein 
(’98) proposes the term “ chromatosphere.” 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


539 


plasmatiques ” of Carnoy, and the “ Plasmosomata ” of the 
other observers correspond to true nucleoli in the sense in 
which this term should be used. The existence of Carnoy’s 
“ nucleoles mixtes ” and “ nucleoles-noyaux ” in cells of Metazoa 
appears to be doubtful. List (’96) considers that the paranu- 
cleoli of the egg cells and the nucleoli of the somatic cells 
are homologous, but that the nucleolus proper of the ova 
is different from both; but the chemical differences which he 
finds between these kinds of nucleoli do not prove that they 
are morphologically distinct structures. 


Appendix to the Literature Reviews. 

Siebold (’39) noticed “ in den Eiern von Plumatella campanu- 
lata Lam. . . . ein deutliches Keimblaschen mit gedoppeltem 
Keimflecke.” 

Koelliker (’43) concludes : “ Es bestande . . . das Ei aus 
einer primitiven Zelle, dem Keimblaschen, die sich um einen 
Kern, den Keimfleck, gebildet, und um die sich nachher Korner 
und eine secundare Zelle, die Dotterhaut, gelegt hatte.” 

Auerbach (’74a) was the first to emphasize and prove clearly 
that the number of nucleoli is usually quite large, and that they 
are frequently irregular in form (before this time it was generally 
assumed that the usual number of nucleoli was one or two). 
The nucleus is filled with “ Grundsubstanz ” (the “ Zellsaft ” of 
Kolliker) and “ Zwischenkornchen ” ; the latter are distin¬ 
guishable from the nucleoli by their smaller size and different 
refraction. He explains the clear zone around the nucleolus 
and the “ Kernkorperchenkreis ” of Eimer by the action of a 
repulsive force on the part of the nucleolus and of the nuclear 
membrane. He distinguishes several successive stages of the 
nucleus with regard to the number of the nucleoli : enucleolar 
nuclei, at an early embryonal stage ; paucinucleolar nuclei, 
with one or two nucleoli ; pliirinucleolar nuclei, with two to four ; 
and multinucleolar , with more than four. “ Die Zahl der Kern- 
korperchen in einem Kerne betragt 1 - 16 , und in extremen 
Fallen selbst noch viel mehr, bis fiber 190 . Und zwar ist nur 
eine kleine Minderheit aller Kerne durch den Gehalt von nur 


540 


MONTGOMERY. 


[Vol. XV. 


einem oder zwei Nucleoli ausgezeichnet.” He gives a large 
series of data on the number and size of nucleoli in embryonal 
and adult cells of vertebrates and Musca. The enucleolar con¬ 
dition is characteristic for embryonal cells; later a nucleolus 
makes its appearance in the center of the nucleus, though its 
substance is probably derived from the cytoplasm ; new nucleoli 
are formed by successive divisions of the first one. In Teleostii 
the nuclei have fewer nucleoli than those of Amphibia , and those 
of Reptilia fewer than those of Mammalia; from which is con¬ 
cluded that the number increases in advancing phylogeny as in 
the ontogeny. “Je schneller und absolut bedeutender das 
Wachsthum der Zellen ist, desto mehr scheint auch die Ten- 
denz zur Vervielfaltigung der Kernkorperchen obzuwalten.” 
The nuclei of the stomach mucosa of Rana are multinucleolar 
in summer and autumn, while after hibernation they contain 
only one to four nucleoli, which may be due to a process of 
fusion. The substance of nucleoli is similar to that of the 
cytoplasm in structure, capability of movements and of produc¬ 
ing vacuoles ; just as the nucleus is first formed as a vacuole in 
the cytoplasm, so in the substance of a nucleolus (which is 
cytoplasmic in origin) a vacuole is formed which has the same 
relation to the nucleolus as the nucleus has to the cell ; “ bei 
dieser Betrachtungsweise erscheint demnach der Zellkern als 
ein hohler Brutraum, bestimmt, eine junge Zellenbrutin sich zu 
entwickeln, die Nucleoli aber als wahrhaft endogen entstandene 
Tochterzellen.” In higher animals all nucleoli do not become 
daughter-cells, but fulfill some new function ; “ und so werden 
wir auch die urspriingliche Bedeutung der Nucleoli als Fort- 
pflanzungszellen nicht fur ganz unmoglich halten diirfen, wenn 
wir auch auf der anderen Seite nicht zweifeln konnen, dass sie 
in den meisten Kernen der hoheren Organismen ganz andere 
Aufgaben zu erfiillen haben miissen.” 

Auerbach (’74b) studied in life the fecundation and cleavage 
of Strongylus and Ascaris. A short time after the appearance 
of the two copulation nuclei in the ovum, arise in each from 
one to five nucleoli ; “ wenn eine Mehrzahl sich einfindet, so 
kommen sie nicht alle gleichzeitig, sondern eines nach dem 
anderen, in Intervallen von einer halben bis zu einigen Minuten 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


541 


zum Vorschein, und zwar in unregelmassigen, oft betrachtlichen 
Entfernungen von einander.” When the nuclei wander towards 
one another the nucleoli move about, “ indem sie innerhalb 
des Kernraums allerlei gerade, zickzackformige, bogenformige, 
Bahnen durchlaufen, mit einer vergleichsweise erheblichen 
Geschwindigkeit, so dass zuweilen in weniger als einer Minute 
Strecken von der Lange des Kern-Durchmessers zuriickgelegt 
werden ”; during these movements the nucleoli remain perfectly 
spherical. When the copulation nuclei are apposed the nucleoli 
in them suddenly disappear, and the mode of this disappearance 
was determined in one case, though it is exceedingly rapid ; “ das 
Kiigelchen wurde allmahlich blasser und etwas grosser und fuhr 
dann plotzlich auseinander, ein Wolkchen bildend, welches einen 
Augenblick darauf nicht mehr zu sehen war.” The nucleoli 
reappear in the resting nuclei, and in the successive genera¬ 
tions up to the eight-cell stage have the same cycle of changes, 
except that in each generation they are somewhat larger than 
in the preceding. These nucleoli are formed independently of 
one another. By the re-formation of the nuclear vacuole a 
number of cytoplasmic granules pass into the cavity of the 
nucleus, and there fuse to form the nucleoli. 

Reinhard (’ 82 , cited by Braem, ’97) describes in the egg of 
Plumatella different stages of the nucleoli, which may be single, 
double, or even trilobular. 

Wistar Institute of Anatomy and Biology, 

Philadelphia, February 3, 1897. 


542 


MONTGOMERY. 


[Vol. XV. 


LITERATURE LIST. 

(An asterisk marks those papers which I have not seen.) 

’57 Agassiz, L. Contributions to the Natural History of the United 
States of America. First Monograph. Part III. Embryology of 
the Turtle. Boston. 

'74a Auerbach, L. Organologische Studien. Zur Charakteristik und 
Lebensgeschichte der Zellkerne. Heft i. Breslau. 

’74b Auerbach, L. Organologische Studien. Zur Charakteristik und 
Lebensgeschichte der Zellkerne. Heft 2. Breslau. 

’76 Auerbach, L. Zelle und Zellkern. Cohn's Beitr. z. Biol, der Pfian- 
zen. 2. 

’90 Auerbach, L. Zur Kenntniss der tierischen Zellen. I. Sitzungsber. 
d. preuss. Akad. d. IViss. Berlin. 

’96 Auerbach, L. Untersuchungen liber die Spermatogenese von Palu- 
dina vivipara. Jena. Zeit.f. Naturw. 30. 

’84 Ayers, H. On the Development of Oecanthus niveus, etc. Me?n. 
Boston Soc. Nat. Hist. 3. 

’64 Balbiani, E. G. Sur les mouvements qui se manifestent dans la tache 
germinative chez quelques animaux. Compt. Rend, de la Soc. Biol. 
Paris. Sdr. 4. 1. 

* ’65a Balbiani, E. G. Sur les mouvements qui se manifestent dans la 

tache germinative chez quelques animaux. Gazette jnddic. de Paris. 
’65b Balbiani, E. G. Observations sur le r61e du noyau dans les cellules 
animales. Compt. Rend. Acad. Paris. 61. 

’76 Balbiani, E. G. Sur les phenom&nes de la division du noyau cellu- 
laire. Compt. Rend. Acad. Paris. 83. 

’81 Balbiani, E. G. Sur la structure du noyau des cellules salivaires chez 
les larves de Chironomus. Zool. Anz. 4. 

’83 Balbiani, E. G. Sur l’origine des cellules du follicule et du noyau 
vitellin de l’ceuf chez les Gdophiles. Zool. Anz. 6. 

* ’95 Balbiani, E. G. Sur la structure et la division du noyau chez la Spiro- 

chona gemmipara. Ann. de Microgr. 7. 

’85 Bambeke, C. van. Etat actuel de nos connaissances sur la structure 
du noyau cellulaire h l’dtat de repos. Ann. Soc. de Med., Gand. 

’86 Bambeke, C. van. Contribution pour servir a l’histoire de la vdsicule 
germinative. Bull. Acad. roy. Belg. 11. 

’93 Bambeke, C. van. Elimination d’eldments nucleaires dans l’ceuf 
ovarien de Scorpaena scrofa L. Bull. Acad. roy. Belg. 25. 

'97a Bambeke, C. van. L’oocyte de Pholcus phalangioides Fuessl. Com¬ 
munication preliminaires. Bruxelles. 

’97b Bambeke, C. van. L’oocyte de Pholcus phalangioides Fuessl. 
Verh. d. anat. Gesell. Jena. 


No. 2 .] COMPARATIVE CYTOLOGICAL STUDIES . 


’98 Bancroft, F. W. Ovogenesis in Distaplia occidentalis Ritter 
(MSS.), with Remarks on Other Forms. Science . Vol. viii, n. s., 
No. 189 . 

’92 Bannwarth. Untersuchungen iiber die Milz. I. Die Milz der Katze. 
Arch.f mikr. Anat. 38 . 

'94 Belajeff, W. Zur Kenntnis der Karyokinese bei den Pflanzen. 
Flora , Erganzungsband. 

’83 Bemmelen, J. F. van. Untersuchungen iiber den anatomischen und 
histologischen Bau der Brachiopoda Testicardines. Jena. Zeit. 
f Naturw. 16 . 

’69 Beneden, E. van. Sur une nouvelle esp&ce de Grdgarine, etc. Bull. 
Acad. roy. Belg. 28 . 

’75 Beneden, E. van. La maturation de l’oeuf, la fdcondation et les 
premieres phases du ddveloppement embryonnaire des mammifkres, 
etc. Bull. Acad. roy. Belg. 40 . 

'76 Beneden, E. van. Contributions h l’histoire de la vdsicule germina- 
tive et du premier noyau embryonnaire. Bull. Acad. roy. Belg. 41 . 

’80 Beneden, E. van. Contributions k la connaissance de l’ovaire des 
mammif&res. L’ovaire du Vespertilio murinus et du Rhinolophus 
ferrum-equineum. Arch, de Biol. Belg. 1 . 

'83 Beneden, E. van. Recherches sur la maturation de l’ceuf et la fdcon- 
dation. Arch, de Biol. 4 . 

’85 Beneden, E. van and Julin, C. Recherches sur la morphologie des 
Tuniciers. Arch, de Biol. 6 . 

’79 Bergh, R. S. Studien iiber die erste Entwicklung des Eies von Gono- 
thryaea Loveni. Morph. Jahrb. 5 . 

’89 Bergh, R. S. Recherches sur les noyaux de l’Urostyla grandis et de 
l’Urostyla intermedia n. sp. Arch, de Biol. 9 . 

*’94 Bergh, R. S. Vorlesungen iiber die Zelle und die einfachen Gewebe 
des thierischen Korpers. Wiesbaden. 

’42 Bischoff, T. Entwicklungsgeschichte des Kaninchen-Eies. Braun¬ 
schweig. 

*’45 Bischoff. Entwicklungsgeschichte des Hunde-Eies. Muller's Archiv. 

’82 Blochmann, F. Ueber die Entwicklung der Neritina fluviatilis. 
Zeit. f. wiss. Zool. 36 . 

’94 Blochmann, F. Ueber die Kerntheilung bei Euglena. Biol. Cen- 
tralbl. 14 . 

’88 Bohm, A. A. Ueber Reifung und Befruchtung des Eies von Petromy- 
zon Planeri. Arch. f. mikr. Anat. 32 . 

’93 Bohmig, L. Zur feineren Anatomie von Rhodope Veranii Kolliker. 
Arb. d. zool. Inst. Graz. 5 . 

’95 Bohmig, L. Die Turbellaria acoela der Plankton-Expedition. Ergeb. 
d. Plankton-Exped., herausgeg. von V. Hensen. 2 . 

’92 Born, G. Die Reifung des Amphibieneies und die Befruchtung 
unreifer Eier bei Triton taeniatus. Anat. Anz. 7 . 


MONTGOMERY. 


[Vol. XV. 


544 

'94 Born, G. Die Struktur des Keimblaschens im Ovarialei von Triton 
taeniatus. Arch.f. mikr. Anat. 43. 

’97 Bouin, P. Etudes sur Involution normale et l’involution du tube 
sdminifere. Arch, de Anat. Micr. 1. 

’84 Bourne, A. G. Contributions to the Anatomy of the Hirudinea. 

Q. J. M. S. n. s., 24. 

’87 Boveri, T. Zellen-Studien, 1. Jena. 

’88 Boveri, T. Zellen-Studien, 2. Jena. 

'97 Braem, F. Die geschlechtliche Entwickelung von Plumatella fungosa. 

Zoologica, herausgegeb. von Leuckart und Chun. Heft 23. 

'74 Brandt, A. Ueber active Formveranderungen des Kernkorperchens. 
Arch. f. mikr. Anat. 10. 

’78 Brandt, A. Ueber das Ei und seine Befruchtungsstatte, etc. Leipzig. 
'82 Brandt, K. Kerntheilungsvorgange bei einigen Protozoen. Biol. 
Centralbl. 3. 

’89 Brass, A. Die Zelle das Element der organischen Welt. Leipzig. 
’91 Brauer, A. Ueber die Entwicklung von Hydra. Zeit.f.wiss.Zool. 52. 
’92 Brauer, A. Ueber das Ei von Branchipus Grubii v. Dyb. von der 
Bildung bis zur Ablage. Abh. d. Akad. d. Wiss. Berlin. 

’93 Brauer, A. Zur Kenntnis der Spermatogenese von Ascaris megalo- 
cephala. Arch.f. mikr. Anat. 42. 

’94 Brauer, A. Ueber die Encystirung von Actinosphaerium Eichhornii. 
Zeit. f. wiss. Zool. 58. 

’95a Bremer, L. Ueber das Paranuclearkorperchen der gekernten Erythro- 
cyten, etc. Arch.f. mikr. Anat. 45. 

’95b Bremer, L. Die Identitat des Paranuclearkorperchens der gekern¬ 
ten Erythrocyten mit dem Centrosom. Arch.f. mikr. Anat. 46. 
'78 Brock, J. Beitrage zur Anatomie und Histologie der Geschlechts- 
organe der Knochenfische. Morph. Jahrb. 4. 

'93 Brooks, W. K. The Genus Salpa. Mem. Biol. Lab. Johns Hof kins 
Univ. 2. 

*97 Bruyne, C. de. Les “ Cellules doubles.” Verb. d. anat. Gesell. f. 1897. 
Jena. 

’94 Bunting, M. The Origin of the Sex-Cells in Hydractinia and Podo- 
coryne, etc. Journ. of Morph. 9. 

'90 Burger, O. Untersuchungen iiber die Anatomie und Histologie der 
Nemertinen, etc. Zeit. f.wiss. Zool. 50. 

’95 Burger, O. Die Nemertinen des Golfes von Neapel. Fauna und 
Flora des Golfes von Neapel. XXII. Monographic. 

'76 Butschli, O. Studien liber die ersten Entwicklungsvorgange der Ei- 
zelle, die Zelltheilung und die Conjugation der Infusorien. Abh. d. 
Senckenberg. naturforsch. Ges. 10. 

'80 Butschli, O. Protozoa. Bronn's Klassen und Ordnungen. 1. 

'85 Butschli, O. Einige Bemerkungen iiber gewisse Organisationsver- 
haltnisse der Cilioflagellaten und der Noctiluca. Morph. Jahrb. 10. 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 545 

’84 Carnoy, J. B. La biologie cellulaire. Etude comparde de la cellule 
dans les deux regnes. Lierre. 

’85 Carnoy, J. B. La cytodidr&se chez les arthropodes. La Cellule. 1 . 

’86 Carnoy, J. B. La cytodidr£se de l’oeuf, etc. La vdsicule germinative 
et les globules polaire chez quelques Nematodes. La Cellule. 3 . 

’97a Carnoy, J. B. and Lebrun, H. La v^sicule germinative et les glo¬ 
bules polaires chez les Batrachiens. La Cellule. 12 . 

’97b Carnoy, J. B. and Lebrun, H. La fdcondation chez l’Ascaris 
megalocephala. Verh. d. anat. Gesell.f. 1897 . Jena. 

’80 Chun, C. Die Ctenophoren des Golfes von Neapel. Fauna und 
Flora des Golfes von Neapel. 1 . 

’69 Claparede, E. Histologische Untersuchungen liber den Regenwurm 
(Lumbricus terrestris Linnd). Zeit.f. wiss. Zool. 19 . 

’95 Coe, W. R. On the Anatomy of a Species of Nemertean (Cerebratu- 
lus lacteus Verr.), etc. Trans. Connect. Acad. 9 . 

’91 Conklin, E. G. and Brooks, W. K. On the Structure and Develop¬ 
ment of the Gonophores of a certain Siphonophore belonging to the 
Order Auronectae. Johns Hopkins Univ. Circulars. 10 . 

*’84 Courchet, L. Du noyau. Paris(?). 

* ’95 Crety, C. Contributo alia conoscenza dell’ ovo ovarico. Richerche 
Lab. Anat. Roma. 4 . 

’91 Cu£not, L. Etudes morphologiques sur les Echinodermes. Arch.de 
Biol. 11 . 

’95 Cunningham, J. T. Experiments and Observations made at the 
Plymouth Laboratory. 2 . The Development of the Egg in Flat 
Fishes and Pipe Fishes. Journ. Mar. Biol. Assoc. London. 3 . 

’97 Cunningham, J. T. On the Histology of the Ovary and of the Ovarian 
Ova in Certain Marine Fishes. Q. J. M. S. N. s., 40 . 

’91 Davenport, C. B. Observations on Budding in Paludicella and Some 
Other Bryozoa. Bull. Mus. Comp. Zool. Harvard. 22 . 

’89 Davidoff, M. v. Untersuchungen zur Entwicklungsgeschichte der 
Distaplia magnilarva Della Valle, etc. I. Mittheil. d. zool. Stat. v. 
Neapel. 9 . 

’97 Debski, B. Beobachtungen liber Kerntheilung bei Chara fragilis. 
Jahrb. wiss. Botan. 30 . 

’95 Delage, Y. La structure du protoplasma et les theories sur l’herd- 
ditd, etc. Paris. 

’93 Demoor, J. Contribution k l’dtude de la physiologie de la cellule. 
Arch, de Biol. 13 . 

’96 Doflein, F. J. T. Die Eibildung bei Tubularia. Zeit.f. wiss. Zool. 
62 . 

'98 Doflein, F. J. T. Studien zur Naturgeschichte der Protozoen. III. 
Ueber Myxosporidien. SfengeVs Zool. Jahrb. Anat. Abtheil 11 . 

’71 Eimer, T. Die Schnauze des Maulwurfs als Tastwerkzeug. Arch, 
f. mikr. Anat. 7 . 


[VOL. XV. 


546 MONTGOMERY. 

’72 Eimer, T. Untersuchungen iiber die Eier der Reptilien. Arch, f 
mikr. Anat. 8. 

’73 Eimer, T. Zoologische Studien auf Capri. I. Ueber Beroe ovatus. 
Leipzig. 

’75 Eimer, T. Ueber amoboide Bewegungen des Kernkorperchens. 
Arch. f. mikr. Anat. n. 

’78 Eimer, T. Die Medusen physiolpgisch und morphologisch auf ihr 
Nervensystem untersucht. Tubingen. 

* ’90 Eimer, T. Organic Evolution. (English translation.) 

’87 Eisig, H. Monographic der Capitelliden des Golfes von Neapel. 

Fauna und Flora des Golfes von Neapel. 16 . 

*’86 Ellenberger. Beitrag zur Lehre von den Kernkorperchen. Arch. 

f. wiss. u. prakt. Tierheilkunde. 12 . 

’80 Engelmann, T. W. Zur Anatomie und Physiologie der Flimmerzel- 
len. Arch. f. ges. Physiol. 23 . 

’97 Erlanger, R. von. Ueber die Morphologie der Zelle und den 
Mechanismus der Zellteilung. Zool. Centralbl. 4 . 

’97 Fairchild, D. G. Ueber Kerntheilung und Befruchtung bei Basidio- 
bolus ranarum Eidam. Jahrb. f. wiss. Botan. 30 . 

’97 Fauvel, P. Recherches sur les Ampharetiens. Bull. soc. de la France 
et Belgique. Paris. 30 . 

’93 Fick, R. Ueber die Reifung und Befruchtung des Axolotleies. Zeit. 
f. wiss. Zool. 56 . 

’88 Fiedler, K. Ueber Ei- und Samenbildung bei Spongilla fluviatilis. 
Zeit.f. wiss. Zool. 47 . 

’74 Flemming, W. Ueber die ersten Entwicklungserscheinungen am Ei 
der Teichmuschel. Arch. f. mikr. Anat. 10 . 

*’75 Flemming, W. Studien in der Entwicklungsgeschichte der Najaden. 

Sitzber. d. Akad. d. Wiss. Berlin. 71 . 

’80 Flemming, W. Beitrage zur Kenntniss der Zelle und ihrer Lebens- 
erscheinungen. Arch.f. mikr. A fiat. 

’82 Flemming, W. Zellsubstanz, Kern und Zelltheilung. Leipzig. 

’94 Flemming, W. Referat “ Die Zelle.” Merkel und Bonnet's Ergebn. 
d. Anat. u. Entw. Wiesbaden. 3 . 

'97 Flemming, W. Zelle. Merkel und Bonnet's Ergebn. d. Anat. u. 
Entw. 5 . 

’96 Floderus, M. Ueber die Bildung der Follikelhiillen bei den Ascidien. 
Zeit.f. wiss. Zool. 61 . 

’73 Fol, H. Die erste Entwicklung des Geryonideneies. Jena. Zeit.f. 
Naturw. 7 . 

’83a Fol, H. Sur l’oeuf et ses enveloppes chez les Tuniciers. Rec. zool. 
suisse. 1 . 

’83b Fol, H. Sur Porigine des cellules du follicule et de l’ovule chez les 
Ascidies et chez d’autres animaux. Comp. Rend. Acad. Sci. Paris. 
96 . 


No. 2 .] COMPARATIVE CVTOLOGICAL STUDIES. 


547 


’89 Fol, H. Sur l’anatomie microscopique du Dentale. Arch. zool. ex~ 
pir. et gen. 7 . 

*’81 Fontana. Traitd sur le venin de la vip£re. Florence. 

’94 Foot, K. Preliminary Note on the Maturation and Fertilization of 
the Egg of Allolobophora foetida. Journ. of Morph. 9 . 

’87 Fraipont, J. Le genre Polygordius. Fauna und Flora des Golfes 
von Neapel. 

’85 Frenzel, J. Einiges iiber den Mitteldarm der Insecten, sowie iiber 
Epithelregeneration. Arch. f. mikr. Anat. 26 . 

’92 Frenzel, J. Die nucleolare Kernhalbirung. Arch.f.mikr. Anat. 39 . 
’93a Frenzel, J. Ueber einige Argentinische Gregarinen, etc. Jena. 
Zeit.f. Naturw. 27 . 

’93b Frenzel, J. Mitteldarmdriise des Flusskrebses und die amitotische 
Zelltheilung. Arch. f. mikr. Anat. 41 . 

* ’67 Frommann, C. Untersuchungen iiber die normale und pathologische 
Anatomie des Riickenmarks. Jena. 

’84 Frommann, C. Untersuchungen iiber Struktur, Lebenserscheinungen 
und Reaktionen thierischer und pflanzlicher Zellen. Jena. Zeit. f. 
Naturw. 17 . 

*’53 Gabriel. De Cucullani elegantis evolutione. Berolini. 

’95 Galeotti, G. Ueber die Granulationen in den Zellen. Internat. 
Monatsch. f. Anat. u. Physiol. 12 . 

’96 Gerould, J. H. The Anatomy and Histology of Caudina arenata 
Gould. Bull. Mus. Comp. Zool. Harvard Coll. 29 . 

’81 Giard, A. Sur un curieux phdnom&ne de prdfecondation, observd 
chez une Spionide. Co7npt. Rend. Acad. Set. Paris. 93 . 

’93 Gjurasin, S. Ueber die Kerntheilung von Peziza vesiculosa Bullard. 
Ber. deutsch. botan. Gesell. 11 . 

’82 Graff, L. von. Monographic der Turbellarien. I. Rhabdocoelida. 
Leipzig. 

’88 Graff, L. von. Die Annelidengattung Spinther. Zeit.f. wiss. Zool. 
46 . 

’96 Greenwood, M. On Structural Change in the Resting Nuclei of 
Protozoa. Part I. The Macronucleus of Carchesium polypinum. 
Journ. Physiol. London. 20 . 

’83 Gruber, A. Ueber Kerntheilungsvorgange bei einigen Protozoen. 
Journ. Physiol. London. 38 . 

’85 Guignard, L. Nouvelles recherches sur le noyau cellulaire et les 
phdnom^nes de la division communs aux vdgdtaux et aux animaux. 
Ann. Sci. Nat. Paris , Botanie. Sdr. 6 . 20 . 

*’94 Guignard, L. L’origine des spheres directrices. Journ. de Botan. 8 . 
’92a Hacker, V. Die Furchung des Eies von Aequorea Forskalea. 
Arch.f. mikr. Anat. 40 . 

’92b Hacker, V. Die Eibildung bei Cyclops und Canthocamptus. Spen- 
geVs zool. Jahrb. 5 . 


548 MONTGOMERY. [Vol. XV. 

’93a Hacker, V. Das Keimblaschen, seine Elemente und Lageverande- 
rungen. I. Arch, f mikr. Anat. 41. 

'93b Hacker, V. Ueber die Bedeutung des Hauptnucleolus. Vorlaufige 
Mittheilung. Ber. d. naturf Gesell. Freiburg i. B. 7. 

’93c Hacker, V. Das Keimblaschen, seine Elemente und Lageverande- 
rungen. II. Arch./, mikr. Anat. 42. 

’95 Hacker, V. Die Vorstadien der Eireifung. Arch. f. mikr. Anat. 45. 

’96 Hacker, V. Ueber eine neue Form der Geschlechtszellen-Sonderung. 
Ber. d. naturf. Gesell. Freiburg i. B. 10. 

’97a Hacker, V. Die Keimbahn von Cyclops. Neue Beitrage zur 
Kenntniss der Geschlechtszellen-Sonderung. Arch. f. mikr. Anat. 
49 . 

’97b Hacker, V. Ueber weitere Uebereinstimmungen zwischen den Fort- 
pflanzungsvorgangen der Tiere und Pflanzen. Biol. Centralbl. 17. 

’74 Haeckel, E. Anthropogenic oder Entwicklungsgeschichte des Men- 
schen. Leipzig. 

’97 Harper, R. A. Kerntheilung und freie Zellbildung im Ascus. Jahrb. 
f. wiss. Botan. 30. 

’86 Heathcote, F. G. The Early Development of Julus terrestris. 
Q. J. M. S. 26. 

’92 Heidenhain, M. Ueber Kern und Protoplasma. Festschr.f Kb Hi¬ 
ker. Leipzig. 

’95 Held, H. Beitrage zur Struktur der Nervenzellen und ihrer Fort- 
satze. Arch. f. Anat. u. Physiol. 

’87 Henking, H. Untersuchungen liber die Entwicklung der Phalangiden. 
I. Zeit.f. wiss. Zool. 45. 

’90 Henking, H. Untersuchungen liber die ersten Entwicklungsvorgange 
in den Eiern der Insekten. II. Ueber Spermatogenese und deren 
Beziehung zur Entwicklung bei Pyrrhocoris apterus L. Zeit.f. wiss. 
Zool. 51. 

’93 Henneguy, L. F. Le corps vitellin de Balbiani dans l’ceuf des Ver- 
tdbrds. Journ. de PAnat. et de la Physiol. Paris. 29. 

’96 Henneguy, L. F. Lemons sur la cellule. Paris. 

’89a Hermann, F. Beitrage zur Histologie des Hodens. Arch.f. mikr. 
Anat. 34. 

'89b Hermann, F. Die postfotale Histiogenese des Hodens der Maus 
bis zur Pubertat. Arch.f. mikr. Anat. 34. 

’97 Hermann, F. Beitrage zur Kenntniss der Spermatogenese. Arch.f. 
mikr. Anat. 50. 

’95 Herrick, F. H. Movements of the Nucleolus through the Action of 
Gravity. Anat. Anz. 10. 

'76 Hertwig, O. Beitrage zur Kenntniss der Bildung, Befruchtung und 
Theilung des thierischen Eies. I. Morfih. Jahrb. 1. 

’77a Hertwig, O. Beitrage zur Kenntniss der Bildung, Befruchtung und 
Theilung des thierischen Eies. II. Morph. Jahrb. 3. 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 549 

’77b Hertwig, O. Weitere Beitrage zur Kenntniss der Bildung, Befruch- 
tung und Theilung des thierischen Eies. Morph. Jahrb. 3. 

’78a Hertwig, O. Beitrage zur Kenntniss der Bildung, Befruchtung und 
Theilung des thierischen Eies. III. Abtheil. 1. Morph. Jahrb. 4. 
’78b Hertwig, O. Beitrage zur Kenntniss der Bildung, Befruchtung und 
Theilung des thierischen Eies. III. Abtheil. 2. Morph. Jahrb. 4. 
’80 Hertwig, O. Die Chaetognathen. Jena. Zeit.f. Naturw. 14. 

’90 Hertwig, O. Vergleich der Ei- und Samenbildung bei Nematoden. 
Arch. f. mikr. Anat. 

’92 Hertwig, O. Die Zelle und die Gewebe. Jena. 

’76 Hertwig, R. Beitrage zur einheitlichen Auffassung der verschiedenen 
Kernformen. Morph. Jahrb. 2. 

’79 Hertwig, R. Der Organismus der Radiolarien. Jena. 

’84 Hertwig, R. Ueber die Kerntheilung bei Actinosphaerium Eichhornii. 

Jena. Zeit.f. Naturw. 17. n. f., 10. 

’88 Hertwig, R. Ueber Kernstruktur und ihre Bedeutung fur Zelltheilung 
und Befruchtung. Sitzber. d. Gesell. f Morph, u. Physiol. Miin- 
chen. 4. 

’96 Hertwig, R. Ueber die Entwickelung des unbefruchteten Seeigel- 
eies. Festschr. f. Gegenbaur. 2. Leipzig. 

’54 Hessling, T. v. Einige Bemerkungen zu des Herrn Dr. Kebers 
Abhandlung “ Ueber den Eintritt der Samenzellen in das Ei,” etc. 
Zeit. f. wiss. Zool. 5. 

*’59 Hessling, T. v. Die Perlmuschel und ihre Perlen. Leipzig. 

’93 Heuscher, J. Zur Anatomie und Histologie der Proneomenia Slui- 
teri Hubrecht. Jena. Zeit.f. Naturw. 27. 

’84 Heuser. Beobachtungen liber Zellkerntheilung. Botan. Centralbl, 
17- 

’94 Hodge, C. F. A Microscopical Study of the Nerve Cell during Elec¬ 
trical Stimulation. Journ. of Morph. 9. 

’90 Holl, M. Ueber die Reifung der Eizelle des Huhns. Sitzb.d. Akad. 
d. Wiss. v. Wien. 99. 

’93a Holl, M. Ueber die Reifung der Eizelle bei den Saugethieren. 

Verh. d. anat. Gesell ., Jte Versammlung, in Gottingen. Jena. 
*’93b Holl, M. Ueber die Reifung der Eizelle bei den Saugethieren. 

Sit zb. d. Akad. d. Wiss. v. Wien , math.-naturw. Cl. 102. 

’81 Hubrecht, A. A. W. Proneomenia Sluiteri gen. et sp. n. Nieder- 
land. Arch. f. Zool. 1. Supplement. 

'87 Hubrecht, A. A. W. Report on the Nemertea Collected by H. M. S. 

“Challenger.” Challenger Reports. 19. 

’97 Huie, L. Changes in the Cell-organs of Drosera rotundifolia, pro¬ 
duced by Feeding with Egg-albumen. Q. J. M. S. N. s., 39. 

’94 Humphrey, J. E. Nucleolen und Centrosomen. Ber. deutsch. botan. 
Gesell. 12. 

’83 Jensen, O. S. Recherches sur la spermatog^n&se. Arch.de Biol. 4 . 


MONTGOMERY. 


[Vol. XV. 


550 

’84 Jijima, J. Untersuchungen iiber den Bau und die Entwicklungsge- 
schichte der Siisswasserdendrocoelen (Tricladen). Zeit. /. wiss. 
Zool. 40. 

’35 Jones, T. W. On the Ova of Women and Mammiferous Animals, as 
they Exist in the Ovaries before Impregnation. London and Edin¬ 
burgh Philos. Mag. 7. 

’37 Jones, T. W. On the First Changes in the Ova of the Mammifera in 
Consequence of Impregnation, etc. Phil. Trans. 37. 

’93 Jordan, E. O. The Habits and Development of the Newt (Diemyc- 
tylus viridescens). Journ. of Morph. 8. 

*’93a Julin, C. Structure et ddveloppement des glandes sexuelles ; ovo- 
genese, spermatog^nese et fecondation chez Styelopsis grossularia. 
Bull. soc. de la France et Belgique. Paris. 25. 

*’93b Julin, C. Le corps vitellin de Balbiani et les dldments de la cellule 
des Mdtazoaires qui correspondent au macronucleus des Infusoires 
cili£s. Bull. soc. de la France et Belgique. Paris. 

'93 Kaiser, J. E. Die Acanthocephalen und ihre Entwicklung. Chun u. 
Leuckarfs Bibliotheca zoologica. 7. 

’93 Karsten, G. Die Beziehungen der Nucleolen zu den Centrosomen 
bei Psilotum triquetrum. Ber. d. deutsch. botan. Gesell. n. 

’90 Kastschenko, N. Ueber den Reifungsprocess des Selachiereies. 
Zeit.f. wiss. Zool. 50. 

’77 Kennel, J. v. Beitrage zur Kenntnis der Nemertinen. Arbeit, zool. 
Inst. Wurzburg. 4. 

’95 Keuten, J. Die Kerntheilung von Euglena viridis Ehrenberg. Zeit. 
f. wiss. Zool. 60. 

’75 Kidd, P. Observations on Spontaneous Movement of Nucleoli. 
Q. J. M. S. n. s., 15. 

’78 Klein, E. Observations on the Structure of Cells and Nuclei. I. 
Q.J.M.S. 18. 

’79a Klein, E. Observations on the Structure of Cells and Nuclei. II. 
Q.J.M.S. 19. 

’79b Klein, E. Ein Beitrag zur Kenntniss der Struktur des Zellkerns. 

Centralbl. f. d. med. Wiss. Berlin. 17. 

’72 Kleinenberg. Hydra, eine anatomisch-entwicklungsgeschichtliche 
Untersuchung. Leipzig. 

’86 Knappe, E. Das Bidder’sche Organ, etc. Morph. Jahrb. 11. 

’96 Koernicke, M. Untersuchungen iiber die Entstehung und Entwick¬ 
lung der Sexualorgane von Triticum, etc. Verh. d. naturf. Ver. d. 
preuss. Rheinlande. 53. 

’43 Kolliker, A. v. Beitrage zur Entwicklungsgeschichte wirbelloser 
Thiere. I. Ueber die ersten Vorgange im befruchteten Ei. 
Muller's Arch. f. Anat. und Physiol. 

*49 Kolliker, A. v. Beitrage zur Kenntniss niederer Thiere. Zeit. f. 
wiss. Zool. 1. 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 551 

’84 Korschelt, E. Ueber die eigenthiimlichen Bildungen in den Zell- 
kernender Speicheldriisen von Chironomus plumosus. Zool. Anz. 7. 

'89 Korschelt, E. Beitrage zur Morphologie und Physiologie des Zell- 
kernes. Sfengel's Zool. Jahrb. 4. 

’95 Korschelt, E. Ueber Kerntheilung, Eireifung und Befruchtung bei 
Ophryotrocha puerilis. Zeit.f. wiss. Zool. 60. 

’96 Korschelt, E. Ueber die Struktur der Kerne in den Spinndriisen 
der Raupen. Arch, f mikr. Anat. 47. 

’97 Korschelt, E. Ueber den Bau der Kerne in den Spinndriisen der 
Raupen. Arch.f. mikr. Anat. 49. 

* '87 Kosinski. Zur Lehre von verschiedenen Typen der Kernkorperchen 
beim Menschen. (In Russian.) Jeschenedjelnaja klin. Gazeta. 
24. 

*’93 Kosinski. Einige Bemerkungen iiber die safraninophilen Kernkor¬ 
perchen in den Krebszellen. (In Russian.) Arch. d. Lab. f. allg. 
Pathol, d. Univ. v. Warschau. 

'92a Kostanecki, K. v. Ueber Centralspindel-Korperchen bei karyo- 
kinetischer Zelltheilung. Vorlaufige Mittheilung. Merkel und 
Bonnet's Anat. Hefte. 

’92b Kostanecki, K. v. Ueber die Schicksale der Centralspindel bei 
karyokinetischer Zelltheilung. Merkel und Bonnet's Anat. Hefte. 

’98 Kostanecki, K. v. Die Befruchtung des Eies von Myzostoma gla- 
brum. Arch.f. mikr. Anat. 51. 

*’92 Kraepelin, K. DU: deutschen Siisswasserbryozoen. II. Abh. naturw. 
Ver. Hamburg. 10. 

'88 Kultschitzky, N. Ueber die Eireifung und die Befruchtungsvor- 
gange bei Ascaris marginata. Arch.f. mikr. Anat. 32. 

’54 Lacaze-Duthiers. Recherches sur les organes gdnitaux des Acd- 
phales lamellibranches. Ann. sci. nat. Sdr. 4. 2. 

’57 Lacaze-Duthiers. Organisation du dentale. Anti. sci. nat. S6r. 4. 
7- 

’84 Lang, A. Die Polycladen (Seeplanarien) des Golfes von Neapel. 
Fauna und Flora des Golfes von Neafel. 11. 

’93 Lauterborn, R. Ueber Bau und Kerntheilung der Diatomeen. 
Vorlaufige Mittheilung. Verh. d. nat.-med. Ver. v. Heidelberg. 5. 

’95a Lauterborn, R. Protozoenstudien. I. Kern- und Zelltheilung von 
Ceratium hirundinella O. F. M. Zeit.f. wiss. Zool. 59. 

’95b Lauterborn, R. Ueber eine Siisswasserart der Gattung Multicilia 
Cienkowski (M. lacustris n. sp.), etc. Zeit.f. wiss. Zool. 60. 

’96 Lauterborn, R. Untersuchungen iiber Bau, Kerntheilung und Bewe- 
gung der Diatomeen. Leipzig. 

’66 La Valette St. George, v. Ueber den Keimfleck und die Deutung 
der Eitheile. Arch.f. mikr. Anat. 2. 

*’83 La Valette St. George, v. Commentatio de Isopodibus. (Fest¬ 
schrift.) 


552 MONTGOMERY. [Vol. XV. 

’94 Lavdowsky. Von der Entstehung der chromatischen und achromati- 
schen Substanzen in den tierischen und pflanzlichen Zellen. Merkel 
und Bonnet's Anat. Hefte. 

'53 Leuckart. Artikel “ Zeugung ” in Wagner's Handworter buck der 
Physiologie. Bd. iv. 

’49 Leydig, F. Zur Anatomie von Piscicola geometrica, mit theilweiser Ver- 
gleichung anderer einheimischer Hirudineen. Zeit.f wiss. Zool. I. 

’50 Leydig, F. Ueber Paludina vivipara, etc. Zeit.f wiss. Zool. 2. 

’52 Leydig, F. Anatomische Notizen iiber Synapta digitata. Arch, f 
Anat. u. Physiol. 

’55a Leydig, F. Ueber Cyclas cornea Lam. Miiller's Arch. f. Anat. u. 
Physiol. 

’55b Leydig, F. Ueber den Bau und die systematische Stellung der 
Raderthiere. Zeit.f. wiss. Zool. 6. 

’83 Leydig, F. Untersuchungen zur Anatomie und Histologie der Thiere. 
Bonn. 

’85 Leydig, F. Zelle und Gewebe. Bonn. 

’88 Leydig, F. Beitrage zur Kenntniss des thierischen Eies im unbe- 
fruchteten Zustande. Spengel's Zool. Jahrb. 3. 

’97 Lidforss, B. Zur Physiologie des pflanzlichen Zellkernes. Acta 
Univ. Lundensis. 33. 

’96 List, T. Beitrage zur Chemie der Zelle und Gewebe. I. Ueber die 
Farbung thierischer Gewebe mit Berlinerblau. Mittheil. d. zool. 
Stat. v. Neap el. 12. 

’92 Lonnberg, E. Kernstudien. Verb. d. biol. Ver. v. Stockhohn. 4. 

’49 Loven, S. L. Ueber die Entwickelung der Mollusca acephala. (Von 
dem Schwedischen von Creplin iibersetzt.) Arch. f. Naturg. 15. 

'74 Ludwig, H. Ueber die Eibildung im Thierreich. Arbeit, zool.-zoot. 
Inst. Wiirzburg. 1. 

’87a Lukjanow, S. M. Beitrage zur Morphologie der Zelle. I. Arch.f. 
Anat. u. Physiol., physiol. Abtheil., Supplement. 

’87b Lukjanow, S. M. Beitrage zur Morphologie der Zelle. II. Ueber 
die Kerne der glatten Muskelzellen bei Salamandra macul. Arch, 
f. mikr. Anat. 30. 

’88 Lukjanow, S. M. Ueber eine eigenthiimliche Form des Kernkorper- 
chens. Arch.f. mikr. Anat. 32. 

’89 Lukjanow, S. M. Einige Bemerkungen iiber sexuelle Elemente beim 
Spulwurme des Hundes. Arch.f. 7nikr. Anat. 34. 

*’93 Lustig und Galeotti. Cytodologische Studien iiber pathologische 
menschliche Gewebe. Ziegler's Beitr. z. pathol. Anat. 14. 

’91 Macallum, A. B. Contributions to the Morphology and Physiology 
of the Cell. Trans. Canadian Inst. 1. 

’95 Macallum, A. B. On the Distribution of Assimilated Iron Com¬ 
pounds, other than Haemoglobin and Haematins, in Animal and 
Vegetable Cells. Q. J. M. S. n. s., 38. 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


'81 Macfarlane, J. M. The Structure and Division of the Vegetable 
Cell. Trans, of the Botan. Soc. of Edinburgh. 14. 

’85 Macfarlane, J. M. Observations on Vegetable and Animal Cells ; 
Their Structure, Division, and History. I. The Vegetable Cell. 
Trans, of the Roy. Soc. of Edinburgh. 30. 

’92 Macfarlane, J. M. A Comparison of the Minute Structure of Plant 
Hybrids with that of their Parents, and its Bearing on Biological 
Problems. Trans, of the Roy. Soc. of Edinburgh. 37. 

'91 Mann, G. Methods of Differential Nucleolar Staining. Trans, and 
Proc. of the Botan. Soc. of Edinburgh. 19. 

92 Mann, G. The Embryo-sac of Myosurus minimus L.: a Cell Study. 

Trans, and Proc. of the Botan. Soc. of Edinburgh. 19. 

'77 Mark, E. L. Beitrage zur Anatomie und Histologie der Pflanzen- 
lause, insbesondere der Cocciden. Arch. f. mikr. Anat. 13. 

’81 Mark, E. L. Maturation, Fecundation, and Segmentation of Limax 
campestris Binney. Bull. Mus. comp. Zool. Harvard Coll. 6. 

’92 Marshall, W. S. Beitrage zur Kenntniss der Gregarinen. Arch.f. 
Naturg. 59. 

'90 Masius, J. Contribution k l’dtude des Rotateurs. Arch. d. Biol. 10. 
’95 Mead, A. D. Some Observations on Maturation and Fecundation in 
Chaetopterus pergamentaceus Cuvier. Journ. of Morph. 10. 

’89 Melissinos and Nicolaides. Untersuchungen iiber einige intra- 
und extranucleare Gebilde im Pankreas der Saugethiere, etc. Arch, 
f Anat. u. Physioh , physiol. Abtheil. 

’93 Mertens, H. Recherches sur la signification des corps vitellins de 
Balbiani dans l’ovule des mammifkres et les oiseaux. Arch, de 
Biol. 13. 

*’67 Metschnikoff, E. Zur Entwicklungsgeschichte der rothen Blut- 
korperchen. Virchow's Arch. f. Pathol. Anat. 41. 

* ’86 Metschnikoff, E. Embryologische Studien an Medusen. Wien. 
’94 Metzner, R. Beitrage zur Granulalehre. Arch.f. Anat. u. Physiol., 

physiol. Abtheil. 

’86 Meunier, A. Le nucteole des Spirogyra. La Cellule. 3. 

'96 Meves, F. Ueber die Entwicklung der mannlichen Geschlechtszellen 
von Salamandra maculosa. Arch.f. mikr. Anat. 48. 

’97 Meves, F. Zur Struktur der Kerne in den Spinndriisen der Raupen. 
Arch.f. mikr. Anat. 48. 

’96 Michel, A. Des nucldoles composes, notamment dans l’oeuf des ann6 - 
lides. Comp. Rend. Acad. Sci. de Paris. 123. 

* ’97 Michel, A. Sur la composition des nucldoles. Comp. Rend, de la 

Soc. Biol, de Paris. 4. 

'93 Minchin, E. A. Observations on the Gregarines of Holothurians. 
Q. f. M. S. n. s., 34. 

’93 Moll, J. W. Observations on Karyokinesis in Spirogyra. Verb, k. 
Akad. Wetensch. Amsterdam. 1. 


MONTGOMERY. 


[Vol. XV. 


554 

’95 Montgomery, T. H., Jr. Stichostemma eilhardi nov. gen. nov. spec. 
Ein Beitrag zur Kenntnis der Nemertinen. Zeit.f. wiss. Zool. 59. 

’96 Montgomery, T. H., Jr. On the Connective Tissues and Body Cavi¬ 
ties of the Nemerteans, etc. SpengeTs Zool. Jahrb. 10. 

’97a Montgomery, T. H., Jr. Studies on the Elements of the Central 
Nervous System of the Heteronemertini. Journ. of Morph. 13. 

’ 97 b Montgomery, T. H., Jr. Preliminary Note on the Chromatin 
Reduction in the Spermatogenesis of Pentatoma. Zool. Anz. 546. 

’95 Moore, J. E. S. On the Structural Changes in the Reproductive Cells 
during the Spermatogenesis of Elasmobranchs. Q. J. M. S. N. s., 
38 . 

’96 Morgan, T. H. The Production of Artificial Astrosphaeres. Arch, 
f Entwicklungsmech. d. Organismen. 3. 

’97 Mottier, D. M. Beitrage zur Kenntniss der Kerntheilung in den 
Pollenmutterzellen einiger Dikotylen und Monokotylen. Jahrb. f. 
wiss. Botan. 30. 

'94 Murbach, L. Beitrage zur Kenntnis der Anatomie und Entwickelung 
der Nesselorgane der Hydroiden. Arch. f. Naturg. 60. 

’88 Nagel, W. Das menschliche Ei. Arch.f. mikr. Anat. 31. 

’85 Nansen, F. Bidrag til Myzostomernes Anatomi og Histologi. Bergens 
Mus. Aasberetn. 

’82 Nussbaum, M. Ueber den Bau und die Thatigkeit der Driisen. 
Arch.f. mikr. Anat. 21. 

’87 Nussbaum, M. Ueber die Theilbarkeit der lebendigen Materie. II. 
Arch.f. mikr. Anat. 29. 

’83 Ogata, M. Die Veranderung der Pankreaszellen bei der Sekretion. 
Arch.f. Anat. u. Physiol., physiol. Abtheil. 

’97 Pennington, M. E. A Chemico-Physiological Study of Spirogyra 
nitida. Public. Botan. Lab. Univ. of Pennsylvania. 1. 

’90 Pfeffer, W. Zur Kenntniss der Plasmahaut und der Vacuolen, nebst 
Bemerkungen fiber den Aggregatzustand des Protoplasmas und fiber 
osmotische Vorgange. Abh. d. konigl. sachs. Akad. d. JViss., 
math.-physik. Cl. 16. 

’81 Pfitzner, W. Ueber den feineren Bau der bei der Zelltheilung auf- 
tretenden Differenzirungen des Zellkerns. Morph. Jahrb. 7. 

’83 Pfitzner, W. Beitrage zur Lehre vom Bau des Zellkerns und seinen 
Theilungserscheinungen. Arch.f. mikr. Anat. 22. 

’86a Pfitzner, W. Zur morphologischen Bedeutung des Zellkerns. 
Morph. Jahrb. 11. 

’86b Pfitzner, W. Zur Kenntnis der Kerntheilung bei den Protozoen. 
Morph. Jahrb. 11. 

*’86c Pfitzner, W. Zur morphologischen Bedeutung des Zellkerns. 
Virchow's Arch. f. pathol. Anat. 103. 

’95 Pflucke, M. Zur Kenntnis des feineren Baues der Nervenzellen bei 
Wirbellosen. Zeit.f. wiss. Zool. 60. 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 


555 

’63 Pfluger, E. F. W. Ueber die Eierstocke der Saugethiere und des 
Menschen. Leipzig. 

’93 Pizon, A. Histoire de la blastog£n&se chez les Botryllid^s. Ann. 
sci. nat. Paris. 14. 

’86 Platner, G. Zur Bildung der Geschlechtsprodukte bei den Pulmona- 
ten. Arch. f. mikr. A nat. 26. 

’89a Platner, G. Beitrage zur Kenntnis der Zelle und ihrer Theilungs- 
erscheinungen. III. Die direkte Kerntheilung in den Malpighi’schen 
Gefassen der Insekten. Arch. f. mikr. A nat. 33. 

'89b Platner, G. Beitrage zur Kenntnis der Zelle und ihrer Theilungs- 
erscheinungen. IV. Die Entstehung und Bedeutung der Neben- 
kerne im Pankreas, etc. Arch.f. mikr. Anat. 33. 

’89c Platner, G. Beitrage zur Kenntnis der Zelle und ihrer Theilungs- 
erscheinungen. VI. Die Bildung der ersten Richtungsspindel im 
Ei von Aulastomum gulo. Arch. f. mikr. Anat. 33. 

’96 Poirault, G. and Raciborski, M. Ueber konjugate Kerne und kon- 
jugate Kerntheilung. Biol. Centralbl. 16. 

'94 Purcell, F. Ueber den Bau der Phalangidenaugen. Zeit. f. wiss. 
Zool. 58. 

’94 Quatrefage, A. de. Etudes embryog^niques. Mdmoire sur l’em- 
bryogdnie des tarets. Ann. sci. nat. Sdr. 3, 11. 

'85 Rabl, K. Ueber Zelltheilung. I. Morph. Jahrb. 10. 

*’93 Raciborski, M. Zur Morphologie des Zellkerns der keimenden 
Samen. Anz. d. Akad. d. Wiss. v. Krakau. 

*’95 Racovitza, E. G. Notes de biologie. III. Mceurs et reproduction 
de la Rossia macrosoma (D. Ch.). Arch, de zool. exjir. et gin. 
S6r. 3, 2. 

'67 Ransom, W. H. Observations on the Ovum of Osseous Fishes. 
Phil. Trans. London. 157. 

’95a Rath, O. vom. Neue Beitrage zur Frage der Chromatinreduktion 
in der Samen- und Eireife. Arch.f. mikr. Anat. 46. 

’95b Rath, O. vom. Ueber den feineren Bau der Driisenzellen des Kopfes 
von Anilocra mediterranea Leach, etc. Zeit. f. wiss. Zool. 60. 

’82 Rauber. Neue Grundlegungen zur Kenntniss der Zelle. Morph. 
Jahrb. 8. 

* ’41 Reichert. Muller's Arch.f. Anat. und Physiol. 

’83 Rein, G. Beitrage zur Kenntniss der Reifungserscheinungen und 
Befruchtungsvorgange am Saugethierei. Morph. Jahrb. 22. 

* ’82 Reinhard, W. Skizze des Baues und der Entwickelung der Siiss- 

wasserbryozoen. Charkow. (Russian.) 

’94 Reinke, F. Zellstudien. I. Morph. Jahrb. 43. 

’58 Remak, R. Ueber die Theilung der Blutzellen beim Embryo. Mul¬ 
ler's Arch. f. Anat. u. Physiol. 

’93 Repiachoff, W. Zur Spermatologie der Turbellarien. Zeit. f. wiss. 
Zool. 56. 


556 MONTGOMERY. [Vol. XV. 

*’81 Retzius, G. Zur Kenntniss vom Bau des Zellkerns. Biolog. Un - 
tersuch. 

’93 Rhumbler, L. Ueber Entstehung und Bedeutung der in den Kernen 
vieler Protozoen und in Keimblaschen von Metazoen vorkommenden 
Binnenkorper (Nucleolen), etc. Zeit.f. wiss. Zool. 56. 

’95 Rhumbler, L. Beitrage zur Kenntnis der Rhizopoden. IV. Cypho- 
deria margaritacea Schlumb. Zeit. f wiss. Zool. 61. 

’96 Rohde, E. Ganglienzellen und Neuroglia. Ein Kapitel iiber Vermeh- 
rung und Wachsthum von Ganglienzellen. Arch.f. ?nikr. Anat. 47. 

’ 92 a Rosen, F. Beitrage zur Kenntniss der Pflanzenzellen. I. Ueber 
tinktionelle Differenz verschiedener Kernbestandtheile und der Sex- 
ualkerne. Cohn's Beitr. z. Biol. d. PJlanzen. 5. Breslau. 

’ 92 b Rosen, F. Beitrage zur Kenntniss der Pflanzenzellen. II. Studien 
iiber die Kerne und die Membranbildung bei Myxomycetes und Pil- 
zen. Cohn's Beitr. z. Biol. d. PJlanzen. 5. Breslau. 

’95 Rosen, F. Beitrage zur Kenntniss der Pflanzenzellen. III. Kerne 
und Kernkorperchen in meristematischen und sporogenen Geweben. 
Cohn's Beitr. z. Biol. d. PJlanzen. 7. Breslau. 

*’95 Rossi, U. Contributo alio studio della struttura, della maturazione 
e della distruzione delle uova degli Anfibi (Salamandrina perspicil- 
lata e Geotriton fuscus). Pubbl..Istit. Stud. Sup. Firenze, Sez. 
Med. Chir. 

’83 Roule, L. La structure de l’ovaire et la formation des oeufs chez les 
Phalli^siadees. Compt. Rend, de VAcad. de Paris. 96. 

’92 Ruckert, J. Zur Entwicklungsgeschichte des Ovarialeies bei Sela- 
chiern. Anat. Ariz. 7. 

’94 Ruckert, J. Zur Eireifung bei Copepoden. Merkel und Bonnet's 
Anat. Hejte. 12. 

'95 Sacharoff, N. Ueber die Entstehung der eosinophilen Granulationen 
des Blutes. Arch.J. mikr. Anat. 45. 

’95 Sala, L. Experimented Untersuchungen iiber die Reifung und 
Befruchtung der Eier bei Ascaris megalocephala. Arch.J. mikr. 
Anat. 44. 

’88 Sanfelice. Spermatogdn&se des vertdbrds. Arch. ital. de Biol. 10. 

’88 Scharff, R. On the Intra-ovarian Egg of some Osseous Fishes. 
Q.J.M.S. N. s., 28. 

’94 Schaudinn, F. Ueber die Kerntheilung mit nachfolgender Korper- 
theilung bei Amoeba crystalligera Gruber. Sitzber. d. k.-k. Akad. 
d. Wiss. v. Berlin. 38. 

’83 Schauinsland, H. Beitrag zur Kenntniss der Embryonalentwicklung 
der Trematoden. Jena. Zeit. f. Naturw. 16. 

’86 Schauinsland, H. Die embryonale Entwicklung der Bothriocepha- 
len. Jena. Zeit. f. Naturw. 19. 

’88 Schewiakoff, W. Ueber die karyokinetische Kerntheilung der Eu- 
glypha alveolata. Morph. Jahrb. 13. 


No. 2 .] COMPARATIVE CYTOLOGICAL STUDIES. 557. 

’78 Schindler, E. Beitrage zur Kenntniss der Malpighi’schen Gefasse 
der Insecten. Zeit.f. wiss. Zool. 30 . 

’38 Schleiden, M. J. Beitrage zur Phytogenesis. Muller's Arch. f. 
Anat ., Physiol, u. wiss , Med. 

’95 Schloter, G. Zur Morphologie der Zelle. Arch.f. ?nikr. Anat. 44 . 
*’80 Schmitz. Untersuchungen liber die Struktur des Protoplasmas und 
der Zellkerne der Pflanzenzellen. Sitzber. d. Niederrhein. Gesell.f. 
Nattirk. Bonn. 

* Schneider, A. Grdgarines nouvelles ou peu connues. Tablettes 

zoologiques. 1 . 

’75 Schneider, A. Contributions h l’histoire des Grdgarines des invertd- 
brds de Paris et de Roscoff. Arch, de zool. exper. et gin. 4 . 

’83 Schneider, A. Nouvelles observations sur la sporulation du Klossia 
octopiana. Arch, de zool. expir. et gin. Sdr. 2 . 1 . 

’91 Schneider, C. Untersuchungen iiber die Zelle. Arb. zool. Inst. 
Wien. 9 . 

’92 Schottlander, P. Beitrage zur Kenntniss des Zellkerns und der 
Sexualzellen bei Kryptogamen. Cohris Beitr.z. Biol.d. Pflanzen. 6 . 
’65 Schron, O. Ueber das Korn im Keimfleck und in dem Kernkorper- 
chen der Ganglienzellen bei Saugethieren. Untersuch. z. Natur- 
lehre d. Thiere , herausgeg. von J. Moleschott. 9 . 

*’82 Schutz, J. Ueber den Dotterkern, dessen Entstehung, Vorkommen 
und Bedeutung. Dissertation. Bonn. 

'87 Schultze, O. Untersuchungen iiber die Reifung und Befruchtung 
des Amphibieneies. Zeit.f. wiss. Zool. 45 . 

*75 Schulze, F. E. Rhizopodenstudien. V. Arch. f. mikr. Anat. 11 . 
'76 Schwalbe. Bemerkungen iiber die Kerne der Ganglienzellen. Jena. 
Zeit.f. Naturw. 10 . 

* '39 Schwann, T. Mikroskopische Untersuchungen iiber die Ueberein- 

stimmung in Struktur und Wachsthum der Thiere und Pflanzen. Berlin. 
'87 Schwarz, F. Die morphologische und chemische Zusammensetzung 
des Protoplasmas. Cohn's Beitr. z. Biol. d. Pflanzen. 5 . 

’82 Seeliger, O. Eibildung und Knospung von Clavelina lepadiformis. 
Sitzber. d. k.-k. Akad. d. Wiss. v. Wien. 85 . 

* ’80 Shafer, E. A. On the Structure of the Immature Ovarian Ovum 

in the Common Fowl and in the Rabbit. Proc. of the Roy. Soc. of 
London. 30 . 

'90 Sheldon, L. The Maturation of the Ovum in the Cape and New 
Zealand Species of Peripatus. Q. J. M. S. n. s., 30 . 

* ’39 Siebold, C. T. von. Beitrage zur Naturgeschichte der wirbellosen 

Thiere. Neueste Schr. d. 7 iaturf. Ges. Danzig. 3 . 

’90 Smirnow, A. Die Struktur der Nervenzellen im Sympathicus der 
Amphibien. Arch. f. mikr. Anat. 35 . 

’95 Sobotta, J. Die Befruchtung und Furchung des Eies der Maus. 
Arch.f. ?nikr. Anat. 45 . 


558 MONTGOMERY. [Vol. XV. 

'93 Stauffacher, H. Eibildung und Furchung von Cyclas cornea L. 
Jena. Zeit. f. Naturw. n. f., 28. 

’97 Stauffacher, H. Die Urniere bei Cyclas cornea (Lam.). Zeit.f. 
wiss. Zool. 63. 

’88 Steinhaus, J. Les metamorphoses et la gemmation indirecte des 
noyaux dans l’epithdlium intestinal de la Salamandra maculosa. 
Arch, physiol., nor?n. et pathol. Sdr. 4, 2. 

’65 Stepanoff, P. Ueber die Geschlechtsorgane und die Entwicklung 
von Cyclas. Arch, f Naturg. 31. 

’87 Stolnikow. Vorgange in den Leberzellen, insbesondere bei der 
Phosphorvergiftung. Arch./. Anat. u. Physiol ., physiol. Abtheil., 
Supplement. 

* ’80 Strasburger, E. Zellbildung und Zelltheilung. 3te Auflage. Jena. 
’82a Strasburger, E. Ueber den Zellkern. Bo tan. Zeitung. 40. 

’82b Strasburger, E. Ueber den Theilungsvorgang der Zellkerne und 

das Verhaltniss der Kerntheilung zur Zelltheilung. Arch.f. mikr. 
Anat. 21. 

’84 Strasburger, E. Die Controversen der indirekten Kerntheilung. 
Arch.f. ?nikr. Anat. 23. 

’88 Strasburger, E. Ueber Kern- und Zelltheilung im Pflanzenreiche, 
nebst einem Anhang iiber Befruchtung. Jena. 

’93 Strasburger, E. Zu dem jetzigen Stande der Kern- und Zellthei- 
lungsfragen. Anat. Anz. 8. 

’95 Strasburger, E. Karyokinetische Probleme. Jahrb.f. wiss. Botan. 
28. 

’97 Strasburger, E. Ueber Cytoplasmastrukturen, Kern- und Zellthei¬ 
lung. Jahrb.f. wiss. Botan. 30. 

’86 Stuhlmann, F. Die Reifung des Arthropodeneies. Nach Beobach- 
tungen an Insekten, Spinnen, Myriapoden und Peripatus. Ber. d. 
naturforsch. Gesell. v. Freiburg i. B. 1. 

* ’69 Svierczewski. Zur Physiologie des Kerns und Kernkorperchens 

der Nervenzellen des Sympathicus. Centralbl. f med. Wiss. Berlin. 
’97 Swingle, W. T. Zur Kenntniss der Kern- und Zelltheilung bei den 
Sphacelariaceen. Jahrb. f. wiss. Botan. 30. 

’82 Tangl, E. Die Kern- und Zelltheilungen bei der Bildung des Pollens 
von Hemerocallis fulva L. Denkschr. d. Wien. Akad. d. Wiss., 
math.-naturw. Cl. 45. 

* ’94 Toyama. On the Spermatogenesis of the Silkworm. Bull. Coll. 

Agricult. Imp. Univ. Tokio. 2. 

’80 Trinchese, S. I primi momenti dell’ evoluzione nei Mollusci. Atti 
R. Accad. Lincei. Ser. 3, 7. 

*’81 Trinchese, S. Per la fauna marittima italiana. Aeolididae e familie 
affini. Atti R. Accad. Lincei. n. 

*’36 Valentin. Repertorium. T. 1. 

*’39 Valentin. Repertorium. T. 4. 


No. 2 .] COMPARATIVE CYTOLOGICAL STUDIES. 


’95 Van der Stricht, O. La maturation et la fdcondation de l’oeuf 
d’Amphioxus lanceolatus. Bull. Acad. Roy. Belg. Sdr. 3 . 30 . 

'82 Vejdovsky, F. Untersuchungen iiber Anatomie, Physiologic und 
Entwicklung von Sternaspis. Denkschr. d. Wien. Akad. d. Wiss. y 
naturw.-math. Cl. 43 . 

'84 Vejdovsky, F. System und Morphologic der Oligochaeten. Prag. 
’88 Vejdovsky, F. Entwicklungsgeschichtliche Untersuchungen. I. 

Reifung, Befruchtung und Furchung des Rhynchelmis-Eies. Prag. 
’95a Vejdovsky, F. Zur vergleichenden Anatomie der Turbellarien. I. 
Zeit. f. wiss. Zool. 60 . 

’95b Vejdovsky, F. Zur vergleichenden Anatomie der Turbellarien. II. 
Zeit.f. wiss. Zool. 60 . 

’93 Ver Eecke. Modifications de la cellule pancrdatique pendant l’ac- 
tivitd sdcrdtoire. Arch, de Biol. 13 . 

'86 Vigelius, W. J. Zur Ontogenie der marinen Bryozoen. Mittheil. d. 
zool. Slat. v. Neapel. 6 . 

'42 Vogt, C. Embryologie des Salmones. Neuchatel. 

’93 Wager, H. On Nuclear Division in the Hymenomycetes. Ann. of 
Botany. London. 7 . 

’96a Wagner, J. Einige Beobachtungen liber die Spermatogenese bei 
den Spinnen. Zool. Anz. 19 . 

* ’96b Wagner, J. Beitrage zur Kenntnis der Spermatogenese bei den 

Spinnen. Arb. nat. Gesell. St. Petersburg. 26 . 

'35 Wagner, R. Einige Bemerkungen und Fragen fiber das Keimblaschen 
(vesiculagerminativa). Muller's Arch.f. Anat., Physiol. u. wiss. Med. 
’36 Wagner, R. Prodromus historiae generationis hominis atque mam- 
malium. Leipzig. 

’37 Wagner, R. Beitrage zur Geschichte der Zeugung und Entwickelung. 

Abh. d. k. bay. Akad. d. Wiss. Mfinchen. 2 . 

*’39 Wagner, R. Artikel “Ei” in Ersch und Gruber's Encyclofadie. 
1 . Abtheil. 

* ’70 Waldeyer, W. Eierstock und Ei. Ein Beitrag zur Anatomie und 

Entwicklungsgeschichte der Sexualorgane. Leipzig. 

’88 Waldeyer, W. Ueber Karyokinese und ihre Beziehungen zu den 
Befruchtungsvorgangen. Arch. f. 7 nikr. A nat. 32 . 

* ’95 Waldeyer, W. Die neueren Ansichten fiber den Bau und das 

Wesen der Zelle. Deutsch. med. Wochenschrift. 

'93 Wasielevsky. Die Keimzone in den Genitalschlauchen von Ascaris 
megalocephala. Deutsche med. Wochenschrift. 41 . 

’94 Watase, S. On the Nature of Cell-Organization. Biol. Lectures at 
Woods Holl for 1893 . 

*’82 Weismann, A. Beitrage zur Kenntniss der ersten Entwicklungsvor- 
gange im Insektenei. Festgabe fur Henle. Bonn. 

’83 Weismann, A. Die Entstehung der Sexualzellen bei den Hydrome- 
dusen. Jena. 


560 MONTGOMERY. [Vol. XV. 

’89 Weismann und Ischikawa. Ueber die Paracopulationszelle im 
Daphnidenei. Spengel's Zool. Jahrb., Abtheil. f. Anat. 4 . 

’87 Went, F. A. F. C. Beobachtungen iiber Kern- und Zelltheilung. 
Ber. d. deutsch. botan. Gesell. 5 . 

’89 Wheeler, W. M. The Embryology of Blatta Germanica and Dory- 
phora decemlineata. Journ. of Morph. 3 . 

’95 Wheeler, W. M. The Behavior of the Centrosomes in the Fertilized 
Egg of Myzostoma glabrum. Journ. of Morph. 10 . 

’96 Wheeler, W. M. The Sexual Phases of Myzostoma. Mittheil. d. 
zool. S/at. v. Neap el. 12 . 

'97 Wheeler, W. M. The Maturation, Fecundation, and Early Cleav¬ 
age of Myzostoma glabrum Leuckart. Arch, de Biol. 15 . 

'78 Whitman, C. O. The Embryology of Clepsine. Q. J'. M. S. 18 . 

'84 Wielowiejski, H. v. Vorlaufige Bemerkungen iiber die Eizelle. 
Biol. Centralbl. 12 . 

'95 Wilcox, E. V. Spermatogenesis of Caloptenus femur-rubrum and 
Cicada tibicen. Bull. Mus. Comp. Zool. Harvard Univ. 27 . 

’84 Will, L. Ueber die Entstehung des Dotters und der Epithelzellen 
bei den Amphibien und Insekten. Zool. Anz. 7 . 

’85 Will, L. Bildungsgeschichte und morphologischer Werth des Eies 
von Nepa cinerea L. und Notonectaglauca L. Zeit.f. wiss. Zool. 41 . 

'86 Will, L. Oogenetische Studien. I. Die Entstehung des Eies von 
Colymbetes fuscus L. Zeit.f. wiss. Zool. 43 . 

'96 Wilson, E. B. The Cell in Development and Inheritance. New York. 

'94 Wilson, H. V. Observations on the Gemmule and Egg Development 
of Marine Sponges. Journ. of Morph. 9 . 

'92 Wir£n, A. Studien liber die Solenogastres. I. Monographic des 
Chaetoderma nitidulum Lovdn. Kongl. Svenska Vet. A had. Hand¬ 
ling. Stockholm. 24 . 

*’45 Wittich, W. v. Observationes quaedam de aranearum ex ovo evo- 
lutione. Halle. 

’49 Wittich, W. v. Die Entstehung des Arachnideneies im Eierstock, 
etc. Arch.f. Anat. u. Physiol. 

'91 Wolters, M. Die Conjugation und Sporenbildung bei Gregarinen. 
Arch.f. mikr. Anat. 37 . 

’82 Zacharias, E. Ueber den Zellkern. Botan. Zeitung. 40 . 

’85 Zacharias, E. Ueber den Nucleolus. Botan. Zeitung. 43 . 

'93 Zacharias, E. Ueber Chromatophilie. Ber. d. deutsch. botan. 
Gesell. 11 . 

’94 Zacharias, E. Ueber Beziehungen des Zellenwachsthums zur Be- 
schaffenheit des Zellkerns. Ber. d. deutsch. botan. Gesell. 12 . 

* ’93 Zimmermann, A. Beitrage zur Morphologie und Physiologie der 
Pflanzenzellen. Bd. 2 . Tiibingen. 

'96 Zimmermann, A. Die Morphologie und Physiologie des pflanzlichen 
Zellkernes : eine kritische Litteraturstudie. Jena. 


No. 2.] COMPARATIVE CYTOLOGICAL STUDIES. 561 


EXPLANATION OF PLATES XXI-XXX. 

All the figures have been drawn with the aid of the camera lucida, and rep¬ 
resent sections of the structures delineated. Those parts of them which are 
colored represent as accurately as possible the stained preparations from which 
they were copied; in most of the figures only certain portions are colored, the 
other details being filled in with the pencil. In order to show the correct pro¬ 
portionate size of the various cells and nuclei the greater number of the figures 
have been made at a magnification afforded by the homogeneous immersion lens 
T a 2 of Zeiss, with the ocular 4, and unless otherwise specified this may be under¬ 
stood to have been the magnification employed. The following abbreviations have 
been used in the figures: 


c. 

cell. 

N.P. 

metamorphosed portion of 

C.D. 

cell duct. 


nucleus. 

Cen. 

centrosome. 

N. Sap. 

nuclear sap. 

Chr. 

chromatin. 

Nut. Gl. 

nutritive globule. 

Chr. F. 

chromatin filament. 

n. 

nucleolus. 

Chrotn. 

chromosome. 

n. 2. 

nucleolus of the second gen¬ 

C. Mb. 

cell membrane. 


eration. 

C.Sp. 

centrosphere. 

n. D. 

derivatives of the nucleolar 

C. T. N. 

nucleus of connective tissue. 


substance. 

C. T. S. 

connective tissue sheath of 

n. Gr. 

granules of degenerated nu¬ 


the ovarial acinus. 


cleoli. 

Cut. 

cuticula. 

n. Mb. 

nucleolar membrane. 

Cy. PI. 

cytoplasm. 

nn. 

nucleolinus. 

d. C. 

degenerated cells (or cell 

n. Sub. 

nucleolar ground substance. 


substance). 

n. Vac. 

nucleolar vacuole. 

End. PI. 

endoplasm. 

nx. 

nucleolar body of unknown 

Gon. Mb. 

gonadal membrane. 


origin. 

Iv. Mb. 

intravitelline membrane. 

Ps. n. 

pseudonucleolus. 

N. 

nucleus. 

Seer. 

secretion corpuscles. 

N. Bd. 

problematical nuclear body. 

Sp. 

spores. 

N. Fib. 

nuclear fibers. 

Sp.F. 

spindle fibers. 

N. Gr. 

nuclear granules. 

Vac. 

vacuole. 

AT. Mb. 

nuclear membrane. 

Yk. Bl. 

yolk ball. 



Yk. Gl. 

yolk globule. 









COMPARATIVE CYTOLOGICAL STUDIES. 563 


EXPLANATION OF PLATE XXL 

Figs, i-ig : Gregarines from Lineus gesserensis. 

Fig. 1. Smallest individual found (hom. immers., oc. 2. Hermann’s fluid; 
Del. haematoxylin, eosin). 

Fig. 2. Outline of the largest individual. Obj. C., oc. 2. 

PTg. 3. Nucleus (corros. sublimate ; Del. haematoxylin, eosin). 

Fig. 4. Portion of a longitudinal section, though an individual in which 
spores were present (as in 3). 

Fig. 5. The smaller of the two nuclei of Fig. 1. 

Fig. 6. The same gregarine drawn in Fig. 1, but with obj. C., oc. 2 to show 
its relative size to the one of Fig. 2. 

Figs. 7-9. Nuclei (alcohol, sublimate ; Ehrlich-Biondi stain, 3^ hrs.). 

Fig. 10. Nucleus (Flemming’s fluid ; Del. haematoxylin, eosin). 

Fig. 11. Idem (Flemming’s fluid ; Ehrlich-Biondi stain, 23^ hrs.). 

Figs. 12-16. Nuclei (Flemming’s fluid; Del. haematoxylin, eosin). 

Figs. 17-19. Idem (sublimate with 2% acetic acid; aq. sol. methylen blue, 
30 min.; aq. sol. brasilin, 2 l f hrs.). 


Figs. 20-33: Gregarines froiti Carinella annulata (fixation with alcohol, sol. 

sublimate). 

Figs. 20 and 21. Outlines of two individuals. Obj. C., oc. 2. 

Figs. 22-25. Nuclei (Del. haematoxylin, 15 min., alum carmine, 6 hrs.). 

Fig. 26. Nucleus (Ehrlich-Biondi stain, 3 hrs.). 

Figs. 27 and 28. Nuclei (Del. haematoxylin, eosin). 

Fig. 29. Nucleus, only the outlines of the nucleoli drawn. 

Figs. 30-35. Nuclei (as in 26). 

Figs. 36~4g: Nuclei of ganglion cells from the brain of Doto {Fig. 36 , of the 
smallest type of cell; Figs. 37-42, of medium-sized cells; Figs. 43~4g, of 
the colossal cells). 

Fig. 36 (Hermann’s fluid, hrs. ; Lyons blue, 15 min.). 

Figs. 37 and 38 (Hermann’s fluid, Vf hrs.; Ehrl. haematoxylin, hrs., 
eosin, 7 min.). 

Figs. 39 and 40 (alcohol, sol. sublimate; Ehrlich-Biondi stain, 3X hrs.). 

Figs. 41 and 42 (as in 37). 

Fig. 43 (as in 39). 

Fig. 44 (Hermann’s fluid, Vf hrs.; safranin, 92 hrs., gentian violet, 
hrs., orange G., 2 min.). 

Fig. 45 (as in 36). 

Figs. 46 and 47. Two sections of one nucleus (as in 37). 

Fig. 48 (as in 37). 

• Fig. 49 (as in 39). 

Fig. 50. Immature germinal vesicle of Emys (picric acid; Del. haematoxylin). 


564 


MONTGOMERY. 


Figs. Ji-j6 : Nuclei from the muscle cells of the circular viusculature of 

Lineus gesserensis. 

Fig. 51 (alcohol, sol. sublimate; Ehrlich-Biondi stain, 3^ hrs.). 

Fig. 52 (aq. sol. sublimate ; cochineal, 1 hr., Del. haematoxylin, 20 min.). 
Figs. 53 and 54 (aq. sol. sublimate with 2% acetic acid; Ehrl. haematoxylin, 
eosin). 

Fig. 55 (Hermann’s fluid, 30 min. ; Ehrl. haematoxylin, 3 hrs.; eosin, 5 min.). 
Fig. 56 (as in 54). 


.Journal of Morphology I ol.AT. 



Montgomery del. et pmx. 


Pi XXL 



litk. Werner & YArdep Frankfort 0 'M. 





























































* 




















% 


































* 















566 


MONTGOMERY. 


EXPLANATION OF PLATE XXII. 

Figs. 37-63, 63-87: Germinal vesicles of Montagua pilata; Figs. 64, 88, 89 ‘ 

germinal vesicles of Doto. 

Figs. 57-59 (alcohol, sol. sublimate ; Ehrlich-Biondi stain, 3 hrs.). 

Fig. 60 (alcohol, sol. sublimate; Ehrl. haematoxylin, 1 hr. ; eosin, 5 min.). 
Figs. 61-63 (aq. sol. sublimate; Del. haematoxylin, 25 min.; eosin, 5 min.). 
Fig. 64 (alcohol, sol. sublimate; Ehrlich-Biondi stain, hrs.). 

Figs. 65-69 (as in 60). 

Fig. 70 (as in 61). 

Fig. 71 (as in 57). 

Figs. 72-75 (as in 61). 

Fig. 76. Outlines of pseudonucleoli from various ova of one individual (aq, 
sol. sublimate). 

Fig. 77 (as in 61). 

Figs. 78-80 (alcohol, sol. sublimate; Mayer’s acid carmine, 15 min.; nigrosine* 
10 min.). 

Figs. 81-87 (Flemming’s fluid ; Del. haematoxylin, eosin). 

Figs. 88 and 89 (as in 64). 

Figs. 90-97: Nuclei of ganglion cells from the brain of Montagua pilata {Fig. 
93 , from a cell of medium size ; the others from the colossal cells'). 

Figs. 90 and 91 (alcohol, sol. sublimate; Ehrlich-Biondi stain, 3 hrs.). 

Figs. 92-94 (picrosulphuric acid; Del. haematoxylin, 25 min.; eosin, 5 min.). 
Fig. 95 (as in 90). 

Figs. 96 and 97 (Flemming’s fluid; Del. haematoxylin, eosin). 


Figs. 98-101: Blood corpuscles of Doto. 

Fig. 98 (alcohol, sol. sublimate; Ehrlich-Biondi stain, 2% hrs.). 

Figs. 99-101 (Hermann’s fluid, 1 % hrs. ; safranin, 92 hrs.; gentian violet, i^£ 
hrs.; orange G., 2 min.). 


. /onnu/l of'. Vin/tholopj/. lb/.. VI' 


PI.XXII. 



-tgomtrv del. *tpinx 


Jitk WerneriWinter, Frankfort°'N. 
















































































































568 


MONTGOMERY ,i 


EXPLANATION OF PLATE XXIII. 

Fig. 102. Blood corpuscle of Doto (picro-nitro-osmic acid, 35 min.; Del. 
haematoxylin, 30 min.; eosin, 5 min.). 

Figs, 103-133 : Egg development of Tetrastemma catenulatnm. 

Figs. 103-106. Germinal vesicles (aq. sol. sublimate ; Ranvier’s picrocarmine; 
Del. haematoxylin). 

Fig. 107. Portion of a young gonad (as in 103). 

Fig. 108. Immature ovum (as in 103). 

Fig. 109. Germinal vesicle (aq. sol. sublimate; Del. haematoxylin, 25 min.; 
eosin, 5 min.). 

Figs, no and hi. Germinal vesicles (aq. sol. sublimate ; Del. haematoxylin, 
15 min.; eosin, 5 min.). 

Fig. 112. Germinal vesicle with portion of the surrounding cytoplasm (as in 
no). 

Fig. 113. Germinal vesicle (as in no). 

Fig. 114. Idem, with portion of the surrounding cytoplasm (as in no). 

Figs. 115 and 116. Outlines of young ova (as in no). 

Figs. 117-119. Germinal vesicles (as in no). 

Fig. 120. Outline of germinal vesicle, the natural color of the nucleoli shown 
(aq. sol. sublimate). 

Fig. 121. Tangential section of the inner surface of the nuclear membrane, 
the dotted line representing the greatest diameter of the nucleus (as in 109). 

Figs. 122-133. Germinal vesicles (as in 109). 

Figs. 134-136: Outlines of the nuclei of ganglion cells of Piscicola. 

Figs. 134 and 135 (alcohol, sol. sublimate). 

Fig. 136 (Flemming’s fluid, 1 hr.). 


PI xx m. 


h mrru tl of a Morphology I T ol. XI: 



Lxtk. H •: r&Wihtev Frankfort 1 








































































































570 


MONTGOMERY. 


EXPLANATION OF PLATE XXIV. 

Figs. 137-139- Germinal vesicles of Tetrastemma catenulatum, the nucleoli 
omitted in Fig. 139 (aq. sol. sublimate ; Del. haematoxylin, 25 min.; eosin, 5 
min.). 

Figs. 140-158: Egg development of Amp hip or us glutinosus. 

Figs. 140-143. Germinal vesicles (aq. sol. sublimate ; Del. haematoxylin, 20 
min.; eosin, 5 min.). 

Figs. 144-146. Germinal vesicles with surrounding cytoplasm (as in 143). 

Figs. 147-150. Germinal vesicles (as in 143). 

Fig. 151. An abnormally large yolk ball (aq. sol. sublimate; aq. sol. dahlia, 
15 min.; eosin, 5 min.). 

Figs. 152-154. Germinal vesicles (aq. sol. sublimate; haematoxylin, 45 
min. ; ferro-ammonio-sulphate, 45 min.; haematoxylin, 45 min.). 

Fig. 155. Ovum and a part of the gonadal cavity in which yolk balls lie, only 
a portion of the cytoplasm drawn (as in 152). 

Figs. 156-158. Germinal vesicles (as in 154). 

Figs. 159-ijj: Egg development of Linens gesserensis. 

Fig. 159. Nuclei from which the germinal vesicles are derived, from the cyto¬ 
plasm of the gonad (Hermann’s fluid ; safranin, 70 hrs.; gentian violet, 1 hr.; 
orange G., 2 min.). 

Fig. 160. Ovum (as in 159). 

Figs. 161 and 162. Germinal vesicles (as in 159). 

Fig. 163. Group of neighboring nuclei from a gonad, showing mitotic stages 
(aq. sol. sublimate ; Del. haematoxylin, 20 min.; eosin, 5 min.). 

Figs. 164-172. Nuclei from gonads (as in 163). 

Figs. 173-176. Germinal vesicles (as in 163). 

Fig. 177. The largest ovum found, only a part of the cytoplasm drawn (as in 


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57 2 


MONTGOMERY. 


EXPLANATION OF PLATE XXV. 

All the figures refer to the large subcuticular gland cells of Piscicola rapax 
Figs. 178-iqb show stages of the prophase , and Fig. 167 the co77imencement 
of the metaphase of the nucleus. 

Fig. 178. Outline of an immature cell, only a portion of its duct drawn (aq. 
sol. sublimate). 

Figs. 179-181. Immature cells (aq. sol. sublimate; Mayer’s acid carmine, 20 
min. ; nigrosine, 25 min.). 

Fig. 182. Immature nucleus (alcohol, sol. sublimate). 

Fig. 183. Idem (alcohol, sol. sublimate; Ehrlich-Biondi stain, 3 hrs.). 

Figs. 184-189. Nuclei (alcohol, sol. sublimate). 

Figs. 190-194. Stages of the ramification of the nucleus (Flemming’s fluid). 
Figs. 195 and 196. Nuclei at the end of the prophase (alcohol, sol. sublimate). 
Fig. 197. Nucleus at the commencement of the metaphase, discharging its 
nucleoli (Flemming’s fluid). 


.Journal of Morphology Vol.XV. PI. XXV. 
























































































-.? - 















































































































































































































574 


MONTGOMERY. 


EXPLANATION OF PLATE XXVI. 

Figs. 198-203: Large subcuticular gland cells of Piscicola , in stages of the 

metaphase. 

Figs. 198 and 199. Nuclei discharging their nucleoli, only outlines drawn 
(Flemming’s fluid). 

Figs. 200-203. Subsequent stages of the metaphase (as in 198). 

Figs. 204-212 : Egg development of a siphonophore (Rodalia (?) ; all fixed in 
alcohol and stained with Del. haejnatoxylin). 

Fig. 204. Ovum from gonophore, chromatin unstained. Obj. A, oc. 4. 

Fig. 205. Ovum from egg pouch. Obj. C, oc. 4. 

Fig. 206. Germinal vesicle from egg pouch. Obj. C, oc. 4. 

Figs. 207-209. Germinal vesicles from gonophores. Obj. C, oc. 4. 

Fig. 210. Ovum from egg pouch. Obj. C, oc. 4. 

Fig. 211. A large and a small ovum from an egg pouch. Obj. C, oc. 4. 

Fig. 212. Nucleolus from a large ovum of a gonophore. 


.huriutl of jM orphology- / ’>/. XI i 


PI.XXVI. 





lith-.Wenm x WfiiteB FfarMt:'VC 









































































































































































. 























57 6 


MONTGOMERY. 


EXPLANATION OF PLATE XXVII. 

Figs. 213-235: Egg development of Stichostemma eilhardi. 

Fig. 213. Portion of the cell syncytium of an immature gonad (aq. sol. sub¬ 
limate ; aq. sol methylen blue, 5 min.; brasilin, 20 min.). 

Fig. 214. Germinal vesicle (as in 213). 

Fig. 215. Yolk balls (as in 213). 

Fig. 216. Germinal vesicle (as in 213). 

Figs. 217 and 218. Portions of cell syncytia of gonads (as in 213). 

Fig. 219. Germinal vesicle (aq. sol. sublimate; Del. haematoxylin, 15 min.; 
borax carmine, 20 hrs.). 

Fig. 220. Portion of the cell syncytium of a gonad (aq. sol. sublimate ; 
Ehrlich-Biondi stain, 3 hrs.). 

Figs. 221-223. Germinal vesicles (as in 220). 

Figs. 224-227. Idem (Flemming’s fluid; alum carmine, 24 hrs.). 

Fig. 228. Portion of a gonadal syncytium (aq. sol. sublimate ; Del. haema¬ 
toxylin, 15 min.; alum carmine, 22 hrs.). 

P igs. 229 and 230. Germinal vesicles (as in 228). 

Fig. 231. Germinal vesicle (aq. sol. sublimate; Del. haematoxylin, 15 min.; 
alum carmine, 45 hrs.). 

Fig. 232. Idem (aq. sol. sublimate ; picrocarmine, 22 hrs.). 

Fig. 233. Ovum, only a portion of the cytoplasm drawn (Lang’s fluid; alum 
carmine; Del. haematoxylin, 15 min.). 

Fig. 234. Germinal vesicle (aq. sol. sublimate ; Del. haematoxylin, 15 min.; 
alum carmine, 16 hrs.). 

Fig. 235. Germinal vesicle and portion of the cytoplasm (aq. sol. sublimate ; 
Del. haematoxylin, 15 min. ; alum carmine, 24 hrs.). 


Figs. 236-24.8: Egg development of Zygonemertes virescens. 

Figs. 236-241. Germinal vesicles (aq. sol. sublimate ; Del haematoxylin, 20 
min.; eosin, 5 min.). 

Figs. 242 and 243. Idem (aq. sol. sublimate; Ehrlich-Biondi stain, 3 hrs.). 
Figs. 244 and 245. Idem (alcohol, sol. sublimate; picrocarmine; Del. hae¬ 
matoxylin, 20 min.; eosin, 5 min.). 

Fig. 246. Portion of an ovum (as in 242). 

Figs. 247 and 248. Germinal vesicles (as in 242). 


Journal of'Morphology Vol.XV. ' PI. XXVII. 









































578 


MONTGOMERY. 


EXPLANATION OF PLATE XXVIII. 

Figs. 24Q-281: Egg development of Polydora. 

Fig. 249. Nuclear division in a peritoneal cell (alcohol, sol. sublimate; Ehrl. 
haematoxylin, 1 hr.; eosin, 5 min.). 

Figs. 250-254. Free cells of the body cavity (as in 249). 

Fig. 255. Nuclear mitosis (as in 249). 

Figs. 256-259. Mitoses of genital cells (as in 249). 

Fig. 260. Nuclear mitosis (as in 249). 

Figs. 261-266. Immature ova (as in 249). 

Figs. 267 and 268. Germinal vesicles (as in 249). 

Fig. 269. Ovum, only a portion of the cytoplasm drawn (as in 249). 

Figs. 270 and 271. Ova (aq. sol. sublimate with 5% acetic acid ; Ehrlich-Biondi 
stain, 3 hrs.). 

Figs. 272-275. Germinal vesicles (as in 270). 

Figs. 276 and 277. Idem (Perenyi’s fluid, 1 hr.; Ehrlich-Biondi stain, 2^ hrs.). 
Fig. 278. Ovum, only a portion of the cytoplasm drawn (Flemming’s fluid; 
safranin, 70 hrs.; gentian violet, 2 % hrs.; orange G, 2 min.). 

Figs. 279-281. Germinal vesicles (as in 278). 

Figs. 282-2qq: Egg development of Tetrastemma elegans. 

Figs. 282-291. Germinal vesicles (alcohol, sol. sublimate; Del. haematoxy¬ 
lin, 25 min. ; eosin, 5 min.). 

Fig. 292. Ova (Hermann’s fluid; Del. haematoxylin,45 min. ; eosin, 5 min.). 
Figs. 293-299. Germinal vesicles (as in 292). 


Journal of Morphology lol.M: 

240 . 



Chr ■SB *' 


• -g ornery del. et pir.x. 


FI. XXVIII. 




















































■ 




























































































































580 


MONTGOMERY. 


EXPLANATION OF PLATE XXIX. 

Figs. 300-316: Egg development of Piscicola rapax. 

Figs. 300-304. Transverse sections of ovarial acini (alcohol, sol. sublimate; 
Ehrl. haematoxylin, 1 hr.; eosin, 5 min.). 

Figs. 305 and 306. Germinal vesicles (as in 300). 

Fig. 307. Ovum (as in 300). 

Fig. 308. Germinal vesicle (as in 300). 

Figs. 309 and 310. Ova (as in 300). 

Fig. 31 i. First pole spindle in the ovum; only one attraction sphere is drawn, 
and that only partially, the dotted line showing how far its rays extend into the 
cytoplasm (as in 300). 

Figs. 312 and 313. Germinal vesicles (alcohol, sol. sublimate ; Mayer’s acid 
carmine, 20 min.; Lyons blue, 5 min.). 

Fig. 314. Germinal vesicle; the dotted line shows the extension of the 
indented surface of the nucleolus, the unstained small oval space being the exter¬ 
nal opening into it (alcohol, sol. sublimate ; fuchsine, 10 min.). 

Fig. 315. Germinal vesicle (Flemming’s fluid; Ehrl. haematoxylin, 2 hrs.; 
eosin, 10 min.). 

Fig. 316. Ovum with attraction spheres at opposite ends of the nucleus; the 
rays of only one attraction sphere drawn (alcohol, sol. sublimate; Ehrl. haema¬ 
toxylin, 40 min. ; eosin, 5^ min.). 


Figs. 3130-324: Mesenchym cells of Cerebratulus lactens (fixation with alcohol. 

sol. sublimate). 

Fig. 315a. Nucleus (Ehrlich-Biondi stain, 2 hrs.). 

Figs. 316a and 317. Nuclear division in free cells (Ehrl. haematoxylin, 2 
hrs.; eosin, 5 min.). 

Figs. 318 and 319. Nuclei (as in 317). 

Figs. 320-324. Whole cells (as in 317). 

Figs. 323-33 7 •' Nuclei of the muscle cells of the longitudinal musculature of 

Piscicola rapax. 

Fig. 325 (aq. sol. sublimate). 

Fig. 326 (alcohol, sol. sublimate; Ehrl. haematoxylin, 1 hr.; eosin, 5 min.). 
Fig. 327 (Flemming’s fluid, 1 hr.). 

Fig. 328 (aq. sol sublimate). 

Fig. 329 (as in 327). 

Figs. 330 and 331 (as in 328). 

Fig. 332 (as in 327). 

Fig. 333 (Flemming’s fluid, 1 hr.). 

Figs. 334 and 335 (as in 326). 

Figs. 336 and 337 (as in 333). 



lontgom ery del etpmx. 


lith. Werner £ Winter, Frankfcrt m 













































































































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5 82 


MONTGOMERY. 


EXPLANATION OF PLATE XXX. 

All figures refer to the giant cells of Doto. 

Fig. 338. Nucleus (alcohol, sol. sublimate; Ehrlich-Biondi stain, 3^ hrs.). 
Fig. 339. Cell (as in 338; hom. immers., oc. 2). 

Figs. 340 and 341. Nuclei (Hermann’s fluid, 1^ hrs.; safranin, 92 hrs. ; gen¬ 
tian violet, hrs. ; orange G., 2 min.). 

Figs. 342 and 343. Two sections of a single nucleus (as in 338). 

Fig. 344. Nucleus (Hermann’s fluid; Ehrl. haematoxylin, 1 y z hrs.; eosin, 
7 min.). 

Fig. 345. Dividing nucleolus (as in 344). 

Fig. 346. Nucleus (as in 344). 







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